Methods and means for gene silencing

ABSTRACT

Provided are insolated DNA vectors which may be based on Agrobacterium binary vectors and which comprise: (a) a transfer nucleotide sequence comprising (i) a plant active promoter, operably linked to (ii) a recombinant tobacco rattle virus (TRV) nucleic acid which may corresponds to all or part of TRV RNA 1 and which includes: a sequence encoding a TRV trans acting factor, and cis acting elements, which confer on the TRV nucleic acid transcript the ability to replicate in the cytoplasm of a plant cell; and a heterologous nucleotide sequence which is foreign to said virus (which may be a cloning site, or a targeting sequence which is capable of down-regulating expression of a target gene); (b) border sequences which permit the transfer of the transfer nucleotide sequence into a plant cell genome. Preferred vectors include pBTAΔAMPΔ16K (SEQ ID NO: 3) or pBTAΔMP (SEQ ID NO: 2). Also provided are related materials and methods of use of such vectors e.g. to produce a cytoplasmically-replicating RNA which can be used to silence target genes in plants.

TECHNICAL FIELD

[0001] The present invention relates generally to recombinant, replicable, plant-viral based nucleic acid constructs, and methods of use thereof in silencing genes in plants.

PRIOR ART

[0002] In plants, post-transcriptional gene silencing (PTGS) can be manifested as an inhibition of nuclear gene expression after the infection with a virus which has been modified to carry sequence from a nuclear expressed gene (Kjemtrup et al., 1998; Kumagai et al., 1995; Ruiz et al., 1998) PTGS can also be manifested after the insertion of a transgene into the plant genome (Napoli et al., 1990; van der Krol et al., 1990). In 2-20% of the cases, after the plant transformation, the plant shows the loss-of function phenotype for the inserted gene instead of its overexpression (Angell and Baulcombe, 1999 and references therein). In both cases, the loss-of function phenotype is caused by sequence specific RNA degradation.

[0003] When the transgene contains the sequence of a replicating virus carrying sequence from a nuclear expressed gene (amplicon virus), the transgenic plant shows the null phenotype for the homologous plant gene in 100% of the plants expressing the replicating amplicon (Angell and Baulcombe, 1999). This null phenotype is stable and inherited through subsequent generations (Angell and Baulcombe, 1997). Therefore, amplicon technology can be used to identify the function of any gene and at the same time, to have the actual knock-out plant for the gene whose function is being identified.

[0004] There are a number of examples of transgenic plants carrying constructs based on whole viral genomes (Atkinson et al., 1998) Kaido et al., 1995; Mori et al., 1993; Yamaya et al., 1988), but most amplicon work has been done with potato virus X (PVX)(Angell and Baulcombe, 1997; Angell and Baulcombe, 1999). In those studies, PVX amplicon was used to transform Nicotiana species and tomato, and silence a number of endogenous genes in these plants (Angell and Baulcombe, 1999). PVX amplicon plants produced infectious viruses, but without any viral symptoms overlapping the silencing phenotype.

DISCLOSURE OF THE INVENTION

[0005] The present invention is concerned with novel viral amplicon constructs. In preferred forms the present invention is concerned with providing amplicon-based methods and materials which may be more suitable as a tool for functional genomics than those which have been used in the past.

[0006] For instance, PVX amplicon Nicotiana plants may not exhibit silencing of genes expressed in meristems (Angell and Baulcombe, 1999). Additionally, PVX has a relatively narrow spectrum of hosts suggesting that it may be difficult to produce silencing of non-host PVX amplicon plants. For example, Arabidopsis thaliana PVX amplicon plants show only weak silencing (Dalmay et al., 2000) and endogenous genes in particular may be difficult to target (Dalmay, unpublished results).

[0007] The present inventors have developed novel viral amplicon constructs based on tobacco rattle virus (TRV) which seeks to address one or more of the problems of the prior art. TRV is able to invade meristems and has a broad range of hosts, including Arabidopsis thaliana.

[0008] Certain viral expression vectors based on TRV have previously been described in which non-viral proteins were expressed from a sub-genomic promoter (Ratcliff, MacFarlane et al. 1999). In that case the viral RNA was synthesised in vitro and then inoculated into the plant. Other TRV based vectors are disclosed by Hamilton & Baulcombe (1989) J. Gen. Virol 70: 963-968 and Mueller et al (1997) J. Gen. Virol 78: 2085-2088.

[0009] Thus in a first aspect of the present invention there is disclosed a nucleic acid vector which comprises:

[0010] (a) a transfer nucleotide sequence comprising (i) a plant active promoter, operably linked to (ii) a recombinant tobacco rattle virus (TRV) nucleic acid which includes:

[0011] a sequence encoding a TRV trans acting factor, and cis acting elements, which confer on the TRV nucleic acid transcript the ability to replicate in the cytoplasm of a plant cell;

[0012] a heterologous nucleotide sequence which is foreign to said virus;

[0013] (b) border sequences which permit the transfer of the transfer nucleotide sequence into a plant cell nucleus.

[0014] The transfer nucleotide sequence is situated between the border sequences and is capable of being inserted into a plant genome under appropriate conditions. Generally this may be achieved by use of so called “agro-infiltration” which uses Agrobacterium-mediated transient transformation. Briefly, this technique is based on the property of Agrobacterium tumafaciens to transfer a portion of its DNA (“T-DNA”) into a host cell where it may become integrated into nuclear DNA. The T-DNA is defined by left and right border sequences which are around 25 nucleotides in length. In the present invention the border sequences are included around the transfer nucleotide sequence (the T-DNA) with the whole vector being introduced into the plant by agro-infiltration, optionally in the form of a binary-transformation vector.

[0015] By “plant active promoter” is meant a sequence of nucleotides from which transcription may be initiated of DNA operably linked downstream (i.e. in the 3′ direction on the sense strand of double-stranded DNA). “Operably linked” means joined as part of the same nucleic acid molecule, suitably positioned and oriented for transcription to be initiated from the promoter. Nucleic acid operably linked to a promoter is “under transcriptional initiation regulation” of the promoter.

[0016] TRV is a bipartite virus, whose genome is composed of two positive stranded RNAs. RNA 1 carries the genes encoding for the replicase, the movement protein (MP) and a small protein called 16K, the precise function of which is unknown. RNA 2 carries the genes for the coat protein (CP) and two proteins involved in nematode transmission (Hernandez et al., 1995).

[0017] The TRV nucleic acid of the present invention includes cis and trans acting elements permitting replication of said cDNA. The vectors of the present invention will generally not require supplementary proteins and/or nucleic acids from TRV in order to achieve this. For instance the cDNA may correspond to all or part of TRV RNA 1. In preferred forms of the invention minimal amplicon constructs are used wherein genes involved in movement of the virus (e.g. MP) and other genes (e.g. 16K), may be removed, thereby leaving only those genes involved in viral replication i.e. one or more trans factors (replicase genes) and cis factors (5′ and 3′ untranslated regions). Generally the constructs will not encode a coat protein.

[0018] The TRV replicase (as with other defined or recited sequences herein) need not be ‘wild-type’, but may optionally be a variant (e.g. mutant, or other variant, or a substantially homologous derivative) provided that its function (to permit, in conjunction with the cis-elements, replication of the TRV nucleic acid transcript) is not negated. By “Substantially homologous” is meant that the sequence in question shares at least about 70%, or 80% identity, most preferably at least about 90%, 95%, 96%, 97%, 98% or 99% identity with the reference sequence. Identity may be at the nucleotide sequence and/or encoded amino acid sequence level.

[0019] Homology may be over the full-length of the relevant sequence shown herein (e.g. in the sequence Annex) or may be over a part of it. Identity may be determined by the TBLASTN program, of Altschul et al. (1990) J. Mol. Biol. 215: 403-10, or BestFit, which is part of the Wisconsin Package, Version 8, September 1994, (Genetics Computer Group, 575 Science Drive, Madison, Wis., USA, Wisconsin 53711). Preferably sequence comparisons are made using FASTA and FASTP (see Pearson & Lipman, 1988. Methods in Enzymology 183: 63-98). Parameters are preferably set, using the default matrix, as follows:

[0020] Gapopen (penalty for the first residue in a gap): −12 for proteins/−16 for DNA; Gapext (penalty for additional residues in a gap): −2 for proteins/−4 for DNA; KTUP word length: 2 for proteins/6 for DNA.

[0021] The heterologous nucleotide sequence is foreign (non-native) to TRV, which is to say that it does not occur naturally in the TRV viral genome at the position in which it is present in the VIGS vector. The sequence will generally be either a cloning site (to permit the insertion of a desired sequence) or a desired sequence itself. It may be introduced in place of other sequence which has been removed (e.g. MP sequence) or as a fusion with all or part of that sequence.

[0022] Some preferred embodiments of the invention will now be discussed.

[0023] Vector

[0024] Nucleic acid vectors according to the present invention may be provided isolated and/or purified, in substantially pure or homogeneous form, or free or substantially free of other nucleic acid. The term “isolated” encompasses all these possibilities.

[0025] Nucleic acid according to the present invention may be polynucleotides or oligonucleotides, and may include cDNA, RNA, genomic DNA and modified nucleic acids. Where a DNA sequence is specified, e.g. with reference to a figure, unless context requires otherwise the RNA equivalent, with U substituted for T where it occurs, is encompassed.

[0026] Where a nucleic acid (or nucleotide sequence) of the invention is referred to herein, the complement of that nucleic acid (or nucleotide sequence) will also be embraced by the invention. The ‘complement’ in each case is the same length as the reference, but is 100% complementary thereto whereby by each nucleotide is base paired to its counterpart i.e. G to C, and A to T or U.

[0027] Preferably the vector is based on plant binary transformation vector pBINTRA6 (see Materials and Methods below).

[0028] Generally speaking, in the light of the present disclosure, those skilled in the art will be able to construct vectors according to the present invention. Such vectors may include, in addition to the promoter, a suitable terminator or other regulatory sequence such as to define an expression cassette consisting of the recombinant TRV nucleic acid, including the heterologous nucleotide sequence. For further details see, for example, Molecular Cloning: a Laboratory Manual: 2nd edition, Sambrook et al, 1989, Cold Spring Harbor Laboratory Press. Many known techniques and protocols for manipulation of nucleic acid, for example in preparation of nucleic acid constructs, mutagenesis, sequencing, introduction of DNA into cells and gene expression, and analysis of proteins, are described in detail in Protocols in Molecular Biology, Second Edition, Ausubel et al. eds., John Wiley & Sons, 1992. Specific procedures and vectors previously used with wide success upon plants are described by Bevan, Nucl. Acids Res. (1984) 12, 8711-8721), and Guerineau and Mullineaux, (1993) Plant transformation and expression vectors. In: Plant Molecular Biology Labfax (Croy RRD ed) Oxford, BIOS Scientific Publishers, pp 121-148.

[0029] Plant Promoter

[0030] Suitable promoters will be well known to those skilled in the art and will generally either be constitutive or inducible (e.g. developmentally regulated or tissue specific). Preferred examples include the Cauliflower Mosaic Virus 35S (CaMV 35S) gene promoter that is expressed at a high level in virtually all plant tissues. The promoter may in principle be an inducible promoter such as the maize glutathione-S-transferase isoform II (GST-II-27) gene promoter which is activated in response to application of exogenous safener (WO93/01294, ICI Ltd). The GST-II-27 gene promoter has been shown to be induced by certain chemical compounds which can be applied to growing plants. Another suitable promoter may be the DEX promoter (Plant Journal (1997) 11: 605-612).

[0031] Recombinant TRV Nucleic Acid

[0032] This is preferably based on a modified, reduced, cDNA clone of TRV RNA 1. In the Examples herein the strain used is ppk20. However any appropriate strain, which can give rise to replicating, infectious viral transcripts, could be used (see e.g. Macfarlane, 1999 for further examples).

[0033] Within the cDNA it is preferable that non-essential ORFs or other sequences are deleted, provided that the CDNA can still be used to generate (cytoplasmically) replicating, infectious transcripts. Preferably, where the cDNA is based on TRV RNA1 of ppk20, one or both of the open reading frames (MP and 16K) are deleted to leave only the 5′ and 3′ untranslated regions and the viral gene encoding the replicase. One or more of the deleted ORFs may be replaced by a heterologous nucleotide sequence (positioned between the UTRs so as to ensure it is replicated).

[0034] Preferred vectors include pBTAΔMPΔ16K or pBTAΔMP. The sequences are shown in the Sequence appendixes. Naturally substantially homologous variants of the sequence are also included within the scope of the invention. In particular, vectors derived from pBTAΔMPΔ16K and having the characteristics (described herein) of that vector, are also embraced.

[0035] Heterologous Nucleotide Sequence.

[0036] This can in principle be a single or multiple cloning site (i.e. a short non-coding sequence encoding two, three or more restriction endonuclease target sites) to facilitate the incorporation of a desired nucleotide sequence.

[0037] Generally the sequence will be a “targeting sequence” which corresponds to a sequence in a target gene, either in the sense or anti-sense (complementary) orientation, or a sequence which has sufficient homology to a target sequence for down-regulation of expression of the target gene to occur. Such a targeting sequence may be included in the vector anywhere in the viral cDNA irrespective of the location of any subgenomic promoter (provided it does not interfere with the cis-acting replication elements or the coat protein). Generally speaking it may be preferable for the TRV amplicons of the present invention not to include a subgenomic promoter within or operably linked to the heterologous gene sequence. Such preferred vectors have the advantage that they are more stable (reduced likelihood of self-recombination) that those of the prior art such as those described by Ratcliff, MacFarlane et al. (1999) supra which had more than one subgenomic promoter.

[0038] In general the targeting sequence may be derived from a plant nuclear gene or transgene, or a gene on an extrachromosomal element such as a plastid.

[0039] Amplicon induced PTGS are particularly preferred for investigating gene function in that it can be used to impose an intermediate or a null phenotype for a particular gene, which can provide information about the function of that gene in vivo. In such cases the identity of the targeting gene may not be known, but the methods of the present invention may be used to identify it with a particular phenotype.

[0040] The complete sequence corresponding to the coding sequence (in reverse orientation for anti-sense) need not be used. For example fragments of sufficient length may be used. It is a routine matter for the person skilled in the art to screen fragments of various sizes and from various parts of the coding sequence to optimise the relationship between target and targeting sequence. It may be preferable that there is complete sequence identity between the targeting sequence in the vector and the target sequence in the plant, although total complementarity or similarity of sequence is not essential. One or more nucleotides may differ in the targeting sequence from the target gene. Thus, a targeting sequence employed in a construct in accordance with the present invention may be a wild-type sequence (e.g. gene) selected from those available, or a substantially homologous mutant, derivative, variant or allele, by way of insertion, addition, deletion or substitution of one or more nucleotides, of such a sequence. Such a sequence need not include an open reading frame or specify an RNA that would be translatable. A typical construct may include a sequence wherein the homology (similarity or identity) between the targeting sequence and the sequence within the target gene is greater than: 80, 85, 90 or 95%, and/or a sequence which targets at least the initiating ATG codon of the target gene.

[0041] A further possibility is to target a conserved sequence of a gene, e.g. a sequence that is characteristic of one or more genes in one or more pathogens against which resistance is desired, such as a regulatory sequence. Thus a construct may target a conserved sequence within a target gene group such as to down-regulate expression of one or more members of a target gene group. More than one targeting sequence may be included.

[0042] Target genes include those which confer ‘unwanted’ traits in the plant and which it may therefore be desired to silence using amplicon-induced PTGS. Examples include ripening specific genes in tomato to improve processing and handling characteristics of the harvested fruit; genes involved in pollen formation so that breeders can reproducibly generate male sterile plants for the production of F1 hybrids; genes involved in lignin biosynthesis to improve the quality of paper pulp made from vegetative tissue of the plant; gene silencing of genes involved in flower pigment is production to produce novel flower colours; gene silencing of genes involved in regulatory pathways controlling development or environmental responses to produce plants with novel growth habit or (for example) disease resistance; elimination of toxic secondary metabolites by gene silencing of genes required for toxin production.

[0043] Other aspects of the invention will now be discussed.

[0044] One aspect of the present invention is a process for producing a vector as described above, the process being substantially as set out in the Examples hereinafter. A further aspect is a process for producing a vector as described above, which process comprises the step of cloning a heterologous nucleotide sequence which is a targeting sequence into the vector.

[0045] A further aspect of the present invention includes a method of silencing a target gene in a plant tissue using amplicon induced PTGS which method comprises the steps of introducing a vector as described above into the plant, wherein said vector includes a heterologous nucleotide sequence which is a targeting sequence.

[0046] “Plant tissue” is any tissue of a plant in planta or in culture, including the whole plant an organ thereof, a cutting, or any group of plant cells organised into a structural and functional unit.

[0047] “Silencing” is a term generally used to refer to suppression of expression of a gene. The degree of reduction may be so as to totally abolish production of the encoded gene product, but more usually the abolition of expression is partial, with some degree of expression remaining. The term should not therefore be taken to require complete “silencing” of expression. It is used herein where convenient because those skilled in the art well understand this.

[0048] As discussed above, for introduction into the plant, the vector may be in the form of an Agrobacterium binary vector. The vector is introduced into the plant cell by Agrobacterium-mediated T-DNA transfer, the transfer sequence may be integrated transiently into the plant (cell) genome, and is then transcribed to RNA from the plant promoter. In the published vector of Ratcliff, MacFarlane et al. (1999), the viral cDNA and any cDNA inserted after the sub-genomic promoter was transcribed to infectious RNA in vitro by T7 RNA polymerase and subsequently introduced into the plant.

[0049] Transient Agrobacterium mediated expression in the plant of the vector is the preferred means of introducing the vector.

[0050] Any appropriate method of plant transformation may be used to generate plant cells containing a construct within the genome in accordance with the present invention. Following transformation, plants may be regenerated from transformed plant cells and tissue.

[0051] Successfully transformed cells and/or plants, i.e. with the construct incorporated into their genome, may be selected following introduction of the nucleic acid into plant cells, optionally followed by regeneration into a plant, e.g. using one or more marker genes such as antibiotic resistance.

[0052] Plants transformed with the DNA segment containing the sequence may be produced by standard techniques which are already known for the genetic manipulation of plants. DNA can be transformed into plant cells using any suitable technology, such as a disarmed Ti-plasmid vector carried by Agrobacterium exploiting its natural gene transfer ability (EP-A-270355, EP-A-0116718, NAR 12 (22) 8711-87215 1984), particle or microprojectile bombardment (U.S. Pat. No. 5,100,792, EP-A-444882, EP-A-434616) microinjection (WO 92/09696, WO 94/00583, EP 331083, EP 175966, Green et al. (1987) Plant Tissue and Cell Culture, Academic Press), electroporation (EP 290395, WO 8706614 Gelvin Debeyser—see attached) other forms of direct DNA uptake (DE 4005152, WO 9012096, U.S. Pat. No. 4,684,611), liposome mediated DNA uptake (e.g. Freeman et al. Plant Cell Physiol. 29: 1353 (1984)), or the vortexing method (e.g. Kindle, PNAS U.S.A. 87: 1228 (1990d). Physical methods for the transformation of plant cells are reviewed in Oard, 1991, Biotech. Adv. 9: 1-11.

[0053] Agrobacterium transformation is widely used by those skilled in the art to transform dicotyledonous species. Production of stable, fertile monocot transgenic plants may be achieved e.g. using the techniques of, or analogous to, Toriyama, et al. (1988) Bio/Technology 6, 1072-1074; Zhang, et al. (1988) Plant Cell Rep. 7, 379-384; Zhang, et al. (1988) Theor Appl Genet 76, 835-840; Shimamoto, et al. (1989) Nature 338, 274-276; Datta, et al. (1990) Bio/Technology 8, 736-740; Christou, et al. (1991) Bio/Technology 9, 957-962; Peng, et al. (1991) International Rice Research Institute, Manila, Philippines 563-574; Cao, et al. (1992) Plant Cell Rep. 11, 585-591; Li, et al. (1993) Plant Cell Rep. 12, 250-255; Rathore, et al. (1993) Plant Molecular Biology 21, 871-884; Fromm, et al. (1990) Bio/Technology 8, 833-839; Gordon-Kamm, et al. (1990) Plant Cell 2, 603-618; D'Halluin, et al. (1992) Plant Cell 4, 1495-1505; Walters, et al. (1992) Plant Molecular Biology 18, 189-200; Koziel, et al. (1993) Biotechnology 11, 194-200; Vasil, I. K. (1994) Plant Molecular Biology 25, 925-937; Weeks, et al. (1993) Plant Physiology 102, 1077-1084; Somers, et al. (1992) Bio/Technology 10, 1589-1594; W092/14828). In particular, Agrobacterium mediated transformation is now emerging also as an highly efficient transformation method in monocots (Hiei et al. (1994) The Plant Journal 6, 271-282).

[0054] The generation of fertile transgenic plants has been achieved in the cereals rice, maize, wheat, oat, and barley (reviewed in Shimamoto, K. (1994) Current Opinion in Biotechnology 5, 158-162; Vasil, et al. (1992) Bio/Technology 10, 667-674; Vain et al., 1995, Biotechnology Advances 13 (4): 653-671; Vasil, 1996, Nature Biotechnology 14 page 702).

[0055] Microprojectile bombardment, electroporation and direct DNA uptake are preferred where Agrobacterium is inefficient or ineffective. Alternatively, a combination of different techniques may be employed to enhance the efficiency of the transformation process, eg bombardment with Agrobacterium coated microparticles (EP-A-486234) or microprojectile bombardment to induce wounding followed by co-cultivation with Agrobacterium (EP-A-486233).

[0056] Following transformation, a plant may be regenerated, e.g. from single cells, callus tissue or leaf discs, as is standard in the art. Almost any plant can be entirely regenerated from cells, tissues and organs of the plant. Available techniques are reviewd in Vasil et al., Cell Culture and Somatic Cel Genetics of Plants, Vol I, II and III, Laboratory Procedures and Their Applications, Academic Press, 1984, and Weissbach and Weissbach, Methods for Plant Molecular Biology, Academic Press, 1989.

[0057] The particular choice of a transformation technology will be determined by its efficiency to transform certain plant species as well as the experience and preference of the person practising the invention with a particular methodology of choice. It will be apparent to the skilled person that the particular choice of a transformation system to introduce nucleic acid into plant cells is not essential to or a limitation of the invention, nor is the choice of technique for plant regeneration.

[0058] The present invention may particularly be applied in plants which are natural hosts (compatible with) TRV. By “compatible” is meant capable of operating with the other components of a system, in this case TRV must be capable of replicating in the plant in question. These include Arabidopsis thaliana. Others include (but are not limited to) Allium cepa; Amaranthus caudatus; Amaranthus retroflexus; Antirrhinum majus; snap-dragon; Arachis hypogaea; Avena sativa; Bellis perennis; Beta vulgaris; Brassica campestris; Brassica campestris ssp. napus; Brassica campestris ssp. pekinensis; Brassica juncea; Calendula officinalis; Capsella bursa-pastoris; Capsicum annuum; Catharanthus roseus; Cheiranthus cheiri; Chenopodium album; Chenopodium amaranticolor; Chenopodium foetidum; Chenopodium quinoa; Coriandrum sativum; Cucumis melo; Cucumis sativus; Glycine max; Gomphrena globosa; Gypsophila elegans; Helianthus annuus; Hyacinthus; Hyoscyamus niger; Lactuca sativa; Lathyrus odoratus; Linum usitatissimum; Lobelia erinus; Lupinus mutabilis; Lycopersicon esculentum; Lycopersicon pimpinellifolium; Melilotus albus; Momordica balsamina; Myosotis sylvatica; Narcissus pseudonarcissus; Nicandra physalodes; Nicotiana benthamiana; Nicotiana clevelandii; Nicotiana glutinosa; Nicotiana rustica; Nicotiana sylvestris; Nicotiana tabacum; Nicotiana edwardsonii; Ocimum basilicum; Petunia hybrida; Phaseolus vulgaris; Phytolacca americana; Pisum sativum; Raphanus sativus; Ricinus communis; Salvia splendens; Senecio vulgaris; Solanum melongena; Solanum nigrum; Solanum tuberosum; Spinacia oleracea; Stellaria media; Trifolium pratense; Trifolium repens; Tropaeolum majus; Tulipa; Vicia faba; Vicia villosa; Viola arvensis.

[0059] A further aspect of the present invention provides a method which includes causing or allowing transcription from a construct as disclosed within the genome of a plant cell to produce a cytoplasmically-replicating RNA.

[0060] A further aspect of the present invention provides a method of reducing or suppressing or lowering the level of a target gene in a plant cell, the method including causing or allowing transcription from a vector as disclosed above.

[0061] In preferred forms the present invention is concerned with providing amplicon-based methods are useful in functional genomics.

[0062] Thus in one aspect of the present invention, the target gene may be of unknown phenotype, in which case the TRV amplicon system may be employed to analyse the phenotype by generating a widespread null (or nearly null) phenotype. The target gene may be essential, which is to say that the null phenotype is lethal to the cell or tissue in question.

[0063] This aspect of the invention may comprise a method of characterizing a target gene comprising the steps of:

[0064] (a) silencing the target gene in a part or at a certain development stage of the plant using the TRV amplicon system described above,

[0065] (b) observing the phenotype of the part of the plant in which, or when, the target gene has been silenced.

[0066] Generally the observation will be contrasted with a plant wherein the target gene is being expressed in order to characterise (i.e. establish one or more phenotypic characteristics of) the gene.

[0067] The potential advantage of the TRV system over certain prior art constructs is discussed above. There are also several advantages of the current method over alternative methods in which the targeted gene is inactivated by insertional or other mutagenic procedures. The advantage over mutagenic procedures applies when there is more than one homologous gene carrying out the role of the target gene. Mutagenic procedures will not normally reveal a phenotype in that situation. A second situation where the current invention has advantage over both mutagenic and unregulated gene silencing procedures applies when the target gene has a lethal phenotype. The controllable attribute of the gene silencing will allow the phenotype of such genes to be investigated and exploited more efficiently than using the alternative methods available prior to the disclosure of the current invention.

[0068] Nor, for the identification of endogenous genes, would it be necessary to try and generate a transgenic plant in which gene silencing is already activated to observe the effect, although transgenic plants may be used if required.

[0069] In a further aspect there is disclosed a method of altering the phenotype of a plant comprising use of the silencing method discussed above. Traits for which it may be desirable to change the phenotype include the following: colour; disease or pest resistance; ripening potential; male sterility.

[0070] In a further aspect of the present invention there is disclosed a kit comprising a vector as described above.

[0071] In a further aspect of the present invention there is disclosed a host cell including a vector according to the present invention. These may be plant cells, or may be microbial (particularly bacterial and especially Agrobacterium) cells. Use of vector as described above in the transformation (stable or transient) of a plant is also embraced by the invention. The host cell may have incorporated into its genome a construct as described above.

[0072] In a further aspect there is disclosed a plant, or plant tissue, stably or transiently transformed by, a vector of the present invention. Thus in addition to a plant, the present invention provides any clone of such a plant, selfed or hybrid progeny and other descendants, and any part of any of these, such as propagules, (any part which may be used in reproduction or propagation, sexual or asexual, including cuttings, seed and so on). Plant extracts and derivatives are also provided. In each case the material will include, or be transformed by, the vector of the present invention.

[0073] The invention will now be further described with reference to the following non-limiting Figures and Examples. Other embodiments of the invention will occur to those skilled in the art in the light of these.

SEQUENCES, FIGURES AND EXAMPLES SEQUENCE APPENDICES

[0074] The sequence of pBTAΔMP is given in full, including vector backbone. For the sequences of pBTAΔMPΔ16K, pBTAΔREPΔMP, and pBTAΔREPΔMPΔ16K, the vector backbone is not given.

[0075] 1. pBINTRA6 (SEQ ID NO: 1)

[0076] 2. pBTAΔMP (SEQ ID NO: 2)

[0077] 3. pBTAΔMPΔ16K (SEQ ID NO: 3)

[0078] 4. pBTAΔREPΔMP (SEQ ID NO: 4)

[0079] 5. pBTAΔREPΔMPΔ16K (SEQ ID NO: 5)

[0080] 6. A. thaliana partial cDNA sequence sulphur gene (SEQ ID NO: 6)

[0081] 7. A. thaliana partial CDNA sequence RUBISCO small subunit gene (SEQ ID NO: 7)

[0082] 8. A. thaliana partial cDNA sequence LEAFY gene (SEQ ID NO: 8)

[0083] 9. mGFP5 cDNA sequence (SEQ ID NO: 9)

FIGURES

[0084]FIG. 1.

[0085] A; Schematic drawing of TRV RNA1; 5′UTR and 3′UTR are the 5′ and 3′ untranslated regions respectively; Rep 134 K is the 134KDa replicase protein; Rep 194 K is the 194 KDa read-through replicase protein; MP is the movement protein; 16K is the 16 KDa protein. B; The relative positions of the PCR1 and PCR2 cDNA fragments.

[0086]FIG. 2.

[0087] Schematic illustration of the cloning strategy for pBSTRF16.

[0088]FIG. 3.

[0089] Schematic representation of the cloning strategy for the introduction of intron 3 from A. thaliana NIA1 gene to TRV RNA 1, to obtain pBSTRA3. INT is NIA1 gene intron 3.

[0090]FIG. 4.

[0091] Schematic representation of pBSTR3′C and pBINTRA6. LB and RB respectively are the left border and right border of pBINTRA6 T-DNA.

[0092]FIG. 5.

[0093] Schematic representation of the construction of pBSTR3′Δ16, plasmid carrying a deletion in the 16K gene. Rep 194K is the 3′ end of viral replicase. MP is the movement protein. 16K is the 16 kD protein gene. 3′ UTR is the 3′ untranslated region. Δ16K is the remaining part of the 16K gene.

[0094]FIG. 6.

[0095] Schematic representation of the construction of pBSTR3′ΔMPΔ16K. AscI, PmlI and PacI are the restriction enzyme sites introduced into the multiple cloning site. ΔMP is the remaining part of the MP.

[0096]FIG. 7.

[0097] Schematic representation of the construction of pBSTR3′ΔMP.

[0098]FIG. 8.

[0099] Schematic representation of the construction of the binary constructs pBTAΔMP and pBTAΔMPΔ16K.

[0100]FIG. 9.

[0101] Construction of negative controls pBTAΔREPΔMP (A) and pBTAΔREPΔMPΔ16K (B). ΔINT is the remaining part of the intron. ΔRep is the remaining part of the viral replicase.

EXAMPLES General Materials and Methods

[0102] Unless stated otherwise, all DNA modifications and digestions were performed using enzymes according to the manufacturers' instructs and following protocols described by Sambrook et al. (Sambrook, Fritsch et al. 1989). Unless stated otherwise, For cloning in E. coli, single or low copy-number vectors were preferred.

[0103] Construction pBINTRA6 and pBSTR3′C

[0104] pBINTRA6 is a full length infectious clone of TRV (strain PPK20) RNA1.

[0105] All the manipulations in TRV RNA1 had to be done first in the plasmid pBSTR3′C because it has more unique sites than pBINTRA6. The vectors were constructed as follows:

[0106] Total RNA was prepared from TRV (strain ppk20) infected N. benthamiana plants as previously described (Devic, Jaegle et al. 1989). Full length cDNA corresponding to TRV RNA1 was prepared from this RNA using Superscript Reverse Transcriptase (Gibco) and the primer TRV2 5′ggggggatccgggcgtaataacgcttacg3′ (SEQ ID NO: 10) which anneals to the 3′ end of TRV RNA1. All primers in this work were derived from the sequence of a closely related TRV strain SYM (Hamilton, Boccara et al. 1987) The full-length cDNA was used as a template for PCR amplification of two overlapping fragments, PCR1 and PCR2, which together cover all of TRV RNA1.

[0107] PCR1, a 3.2 kb fragment, was amplified using Expand HiFi polymerase (Roche). The primers were: TRV1 ′ggggggatccataaaacatttcaatcctttg3′ (SEQ ID NO: 11) (which anneals to positions 1-21 of TRV) and TRV4U 5′ttagcaccagctatctgagcgc3′ (SEQ ID NO: 12) (positions 3168-3189). PCR2, a 4.1 kb product, was also amplified using Expand HiFi polymerase (Roche) and the primers TRV4D 5′gttccaaccagacaaacgtatgg3′ (SEQ ID NO: 13) (positions 2698-2720) and TRV2 (see above).

[0108] PCR1 and PCR2 share a 491 nt overlap in the replicase open reading frame (ORF). The primers TRV1 and TRV2 contain BamHI sites to allow cloning of the full-length product (FIG. 1).

[0109] PCR2 was blunt-ended using T4 DNA polymerase, digested with BamHI, and cloned into the plasmid pBAC/SacB1 (Bendahmane, Kanyuka et al. 1999) which had previously digested with BamHI and EheI to form pBSTR3′ C. The PCR1 fragment was blunted-ended with T4 DNA polymerase and ligated into HpaI digested-pBSTR3′C, to form pBSTRF16. pBSTRF16 therefore contains 302 bp that are duplicated within the replicase ORF (FIG. 2).

[0110] The 302 bp of duplicated replicase sequence was replaced with a 438 bp intron. Intron 3 of Arabidopsis thaliana Col-0 nitrate reductase NIA1 gene (Wilkinson and Crawford 1993) was amplified using the primers AraF and AraR. AraF is 5′CGTATCTTTGCAA TAACAGgtaataatcctctctcttgatatt3′ (SEQ ID NO: 14), where the sequence in upper case corresponds to positions 2826-2845 of TRV RNA1 and the sequence in lower case corresponds to positions 1-24 of the intron. Similarly, AraR is 5′TTAAATTGTCCAAGATCAACct gtttaacacaagtcaacgtc3′ (SEQ ID NO: 15) where the sequence in upper case corresponds to positions 2846-2864 of TRV RNA 1 and the sequence in lower case corresponds to positions 416-438 of the intron. The PCR amplified intron 3 fragment was therefore flanked by the AGGT intron splice-sites, and 19 bp of TRV (exon) sequence (FIG. 3).

[0111] Two TRV-exons (exon 1 and exon 2) that flank the intron insertion site were then PCR amplified. For exon 1 the primers were TRV2D 5′tcgcacaaaaccaaggtgatag3′ (SEQ ID NO: 16) (positions 1772-1793) and Ara5′R 5′ggattatt acCTGTTATTGCAAAGATACGTCTG3′ (SEQ ID NO: 17) where the sequence in lower case corresponds to positions 1-10 of the intron and sequence in upper case corresponds to positions 2822-2845 of TRV RNA1. Exon 1 was amplified as a 1.07 kb fragment from pBSTR16. For exon 2 the primers were Ara3′F 5′tgttaaacagGTTGATC TTGGACAATTTAAGTGC3′ (SEQ ID NO: 18), where the sequence in upper case corresponds to positions 2846-2868 of TRV RNA1 and the sequence in lower case corresponds to positions 428-438 of the intron, and TRV4U (see above). Exon 2 was amplified as a 0.35 kb fragment from PCR 1 (see above). Exon 1, intron3 and exon 2 were all amplifed using Pfu polymerase (Promega). To introduce intron 3 to the TRV RNA 1 genome, chimeric PCR was performed with Pfu polymerase and the primers TRV2D and TRV4U using a mixture of exon 1, intron 3 and exon 2 as template to give a 1.8 kb fragment.

[0112] This 1.8 kb intron-containing-fragment was digested with ApaI and SalI and cloned in pBSTRF16 using ApaI-partial digestion and SalI, thus replacing the region that included duplicated sequence, and forming pBSTRA3 (FIG. 3).

[0113] To transfer the cloned RNA1 to a binary transformation vector, the 7.2 kb fragment corresponding to TRV RNA 1 was released from pBSTRA3 with BamHI and cloned into the BamHI site between the CaMV 35s promoter and the CaMV terminator on the plasmid pBIN61 to form pBINTRA6. pBIN61 is a modified version of the pBIN19 (Frisch, Harris-Haller et al. 1995) binary vector that carries a transcription cassette comprising the CaMV 35S promoter and terminator. To construct the pBIN61 binary vector, the transcription cassette containing the CaMV 35S promoter and terminator was released by digestion with KpnI and XhoI from the plasmid pJIT61 (kindly provided by P. Mullineaux, JIC, Norwich, UK). The transcription cassette was then ligated to the pBIN19 plasmid vector digested with KpnI and SalI to create pBIN61. pBIN61 is a low copy number vector in E. coli (10-15 copies per cell) in which the TRV insert can be stably cloned.

[0114] Agrobacterium strain GV3101 containing pBINTRA6 was infiltrated into N. benthamiana leaves causing a TRV RNA 1 infection. The full sequence of pBINTRA6 is given in the Appendix

[0115] A schematic representation of pBSTR3′C and pBINTRA6 is shown in FIG. 4.

[0116] Construction of pBSTR3′Δ16

[0117] First of all, we had to build a pBSTR3′C derivative bearing a deletion in the 16K sequence so that subsequent manipulations could be done on this derivative, in addition to pBSTR3′C, in order to produce corresponding amplicon vectors with and without the 16K open reading frame.

[0118] Most of the 16K open reading frame was removed from pBSTR3′C via chimeric PCR (Ho et al., 1989; Horton et al., 1989). Two fragments were amplified using as template pBINTRA6 and Pfu I polymerase (Promega). The 5′ PCR fragment was amplified using primers TR5400D: 5′ ttctcaaatctaggggccattg 3′ (SEQ ID NO: 19) corresponding to positions 5381 to 5403 of TRV RNA1 and Δ16R2: 5′ CCGAAAGGAACacttcattcacacaacccttga 3′ (SEQ ID NO: 20), were letters in upper case correspond to positions 6501 to 6511 of TRV RNA1, and letters in lower case correspond to positions 6124 to 6145. This fragment was 0.77 Kb. The 3′ PCR fragment was amplified using primers Δ16F2: 5′ gaatgaagtGTTCCTTTCGGGATTGATCGTT 3′ (SEQ ID NO: 21) where the letters in upper case correspond to positions 6501 to 6522 and the letters in lower case, to positions 6137 to 6145 and TRV2: 5′ggggggatccgggcgtaataacgcttacg3′ (SEQ ID NO: 10) which anneals to the 3′ end of TRV RNA1 (positions 6770-6789). This fragment was 0.3 Kb. Both fragments, therefore, share an overlapping sequence of 20 nucleotides. To produce the deletion, chimeric PCR was performed with Pfu I polymerase and primers TR5400D and TRV2 using a mixture of 5′ and 3′ PCR fragments to give a fragment of 1.07 Kb in which 355 bp from 16Kb open reading frame have been deleted.

[0119] To insert this deletion into pBSTR3′C, the PCR fragment was digested with MluI and SnaBI and inserted in the MluI and SnaBI sites of pBSTR3′C to give the plasmid pBSTR3′Δ16 (FIG. 5).

[0120] Construction of pBTAΔMP and pBTAΔMPΔ16K

[0121] Part of the movement protein sequence was removed from pBINTRA6 and a multiple cloning site was engineered and put in its place via chimeric PCR (FIG. 6).

[0122] To carry out the deletion two fragments were amplified using PfuI polymerase and pBINTRA6 as template. The 5′ PCR fragment was amplified using primers TR4870D: 5′actcactgattgcgtttcctag 3′ (SEQ ID NO: 22) (positions 4848-4869) and ΔMPR: 5′ ttaattaacacgtggcgcgccAGTCTTCTTCTTCAAGGTGACC 3′ (SEQ ID NO: 23), where the sequence in lower case corresponds to the sequence of AscI-PmlI-PacI sites of the engineered polylinker and the sequence in upper case, to positions 5345 to 5366 of TRV RNA1. This fragment was 0.54Kb. The 3′ PCR fragment was amplified using primers ΔMPF: 5′ ggcgcgccacgtgttaattaaCTGATTCGACTAGGCGCCTC 3′ (SEQ ID NO: 24), where the sequence in lower case corresponds to the sequence of AscI-PmlI-PacI sites of the engineered polylinker and the sequence in upper case, to positions 5857 to 5876. and TRV2 (see above). This fragment was 0.96 Kb. Both fragments share a 21 nucleotides fragment corresponding to the engineered polylinker. The actual deletion and introduction of the polylinker was made via chimeric PCR using PfuI polymerase and primers TR4870 and TRV2 and a mixture of both PCR fragments. The product was 1.5 Kb. Then, it was digested with AatII and EheI and introduced into the AatII and EheI sites of pBSTR3′Δ16 to produce pBSTR3′ΔMPΔ16 (FIG. 6).

[0123] To produce the corresponding construct carrying only the deletion in the MP gene and not in the 16K gene, pBSTR3′ΔMPΔ16 was digested with EheI and BamHI to remove a 568 bp fragment including the 16K deletion and replaced by a 923 bp BamHI-EheI fragment from pBSTR3′C carrying the full length 16K gene (FIG. 7).

[0124] To introduce these deletions into the binary construct pBINTRA6, pBSTR3′Δ16 and pBSTR3′ΔMPΔ16 were digested with AvrII and StuI and the fragments containing the deletions were cloned into the AvrII and StuI sites of pBINTRA6 to produce pBTAΔMP and pBTAΔMPΔ16 (FIG. 8)

[0125] Construction of the Negative Control Amplicon Vectors

[0126] The corresponding negative control, non replicative vectors bearing a deletion on the viral replicase gene were constructed by digesting both pBTAΔMP and pBTAΔMPΔ16 with SwaI and HpaI, which have unique sites on these vectors and produce blunt ends. Then the resulting fragment was religated, to produce either pBTAΔRepΔMP or pBTAΔRepΔMPΔ16. Since the SwaI site was inside the intron, these constructs have lost 368 bp of the intron and 40 bp of the replicase. They have also lost one of the intron splicing sites and, therefore, will be unable to splice the intron to produce a native replicase (FIGS. 9A and 9B).

[0127] Construction of Amplicon-Derived Constructs

[0128] Sulphur Constructs

[0129] Sulphur codifies for a magnesium chelatase, an enzyme required for chlorophyll formation. A 944 bp cDNA fragment was amplified from pTV09 plasmid (F. Ratcliff, unpublished), which contained a 1.2 Kb of A. thaliana cDNA sulphur gene (Kjemtrup et al., 1998). It was amplified using Expand HiFi polymerase and the primers SUL1: 5′ ccttggcgcgccttcactctcttctccttcc (SEQ ID NO: 25) and SUL2: 5′ ccccttaattaatctggtcttgaagcttgtcc (SEQ ID NO: 26). SUL1 carries a restriction site for AscI and SUL2, one for PacI to facilitate the insertion of the fragment into the AscI and PacI sites of the multiple cloning site of the amplicon vectors The resulting constructs were pBTAΔMP:S, pBTAΔMPΔ16K:S, pBTAΔREPΔMP:S and pBTAΔREPΔMPΔ16:S. The sequence is given in the Appendix.

[0130] RUBISCO Constructs

[0131] RUBISCO is a gene involved in carbon fixation during photosynthesis. A 469 bp cDNA fragment of the rubisco small sub-unit was PCR amplified from A. thaliana cDNA using Expand HiFi polymerase and the primers aRUB1:5′ ccttggcgcgcctctatgctctcttccgcta (SEQ ID NO: 27) and aRUB2:5′ ccccttaattaatccgatgatcctaatgaaggc (SEQ ID NO: 28). As above, the primers carry restriction sites for AscI and PacI to facilitate the cloning into the corresponding AscI and PacI sites of the multiple cloning site of the amplicon vectors. The resulting constructs were pBTAΔMP:aR, pBTAΔMPΔ16K:aR, pBTAΔREPΔMP:aR and pBTAΔREPΔMPΔ16:aR. The sequence is given in the Appendix.

[0132] LEAFY Constructs

[0133] A 940 bp cDNA fragment of LEAFY, a gene involved in floral development in A. thaliana, was PCR amplified from plasmid pDW122 (Weigel et al., 1992) using Expand HiFi and the primers LEAFY1: 5′ ccttggcgcgccatacggtatacgtttctacac (SEQ ID NO: 29) and LEAFY2: 5′ ccccttaattaaagacggcgtctatatccc (SEQ ID NO: 30). As above, the primers carry restriction sites for AscI and PacI to facilitate the cloning into the corresponding AscI and PacI sites of the multiple cloning site of the amplicon vectors. The resulting constructs were pBTAΔMP:Lfy, pBTAΔMPΔ16K:Lfy, pBTAΔREPΔMP:Lfy and pBTAΔREPΔMPΔ16:Lfy. The sequence is given in the Appendix.

[0134] GFP Constructs p A 790 bp fragment containing the whole coding sequence of mGFP5 was amplified from plasmid CL106 (Haseloff et al., 1997) using Expand HiFi polymerase and the primers 5′GFP: 5′ ggttggcgcgccaatgaagactaatctttttctc (SEQ ID NO: 31) and 3′GFP: 5′ ggggttaattaattagagttcgtcatgtttgta (SEQ ID NO: 32). As above, the primers carry restriction sites for AscI and PacI to facilitate the cloning into the corresponding AscI and PacI sites of the multiple cloning site of the amplicon vectors. This way, the GFP gene is in frame with the first 13 amino acids of the movement protein and will be expressed as a fusion protein. The resulting constructs were pBTAΔMP:GFP, pBTAΔMPΔ16K:GFP, pBTAΔREPΔMP:GFP and pBTAΔREPΔMPΔ16:GFP. The sequence is given in the Appendix.

[0135] To carry out the transient assays in N benthamiana and A. thaliana transformation, all the amplicon derived constructs were introduced into the Agrobacterium strain GV3101.

[0136] Plant Manipulations

[0137] Plant Growth Conditions

[0138] All work involving virus infected material was carried out in containment glasshouses under MAFF license PHF 1420c/1773(December 1996). N. benthamiana and A. thaliana plants were germinated on a 1:1 mixture of JIC compost and peat, then grown individually in pots at 25° C. during the day and 20° C. during the night. Supplementary winter lighting from halogen quartz iodide lamps provided a 16 hour day length.

[0139] Agrobacterium-Infiltration

[0140] Virus infections on N. benthamiana were achieved by Agrobacterium-mediated transient gene expression of infectious constructs from the T-DNA of a binary plasmid (e.g. any of the amplicon constructs). Agrobacterium was grown to saturation in L broth. The culture was then centrifuged and re-suspended in 10 mM MgCl 2, 10 mM MES and 150 mM acetosyringone, and kept at room temperature for 2 hours. The culture was then infiltrated to the underside of a leaf using a 2 ml syringe without a needle.

Example 1

[0141] Transient Assay to Demonstrate Replication

[0142] The ability of the amplicon constructs to replicate in plants is tested on N. benthamiana as follows. Agrobacterium cultures of amplicon constructs carrying the whole GFP gene (pBTAΔMP:GFP, pBTAΔMPΔ16:GFP, pBTAΔREPΔMP:GFP, pBTAΔREPΔMPΔ16:GFP) are infiltrated into all the leaves of N benthamiana plants four weeks old. Ten days after infiltration, the infiltrated patch shows green fluorescence under UV light. Controls unable to replicate do not show green fluorescence in the infiltrated patch. Samples may be taken to confirm the presence of GFP RNA in those plants using northern blotting.

Example 2

[0143] Transient Assay to Demonstrate Silencing

[0144] Ability to produce silencing may be tested on N benthamiana plants as follows. Agrobacterium cultures of amplicon constructs carrying a piece of sulphur gene are infiltrated into all the leaves of N. benthamiana plants four weeks old. Ten days after infiltration the infiltrated patch shows a faint yellow colour typical representing sulphur-silencing in the leaves. Controls unable to replicate, or having weaker promoters, show reduced silencing or no silencing in the infiltrated patch. Samples may be collected to confirm the absence of sulphur RNA from silenced plants using northern blotting.

[0145] Corresponding assays employing GFP-based constructs, in GFP-transgenic N benthamiana plants, may also be performed.

Example 3

[0146] Transformation of A thaliana with Amplicons

[0147] GV3101 Agrobacterium cultures containing individual amplicon constructs were grown in 500 ml L broth in the presence of 50 μg/ml Kanamycin at 28° C. After centrifugation at room temperature the cells were resuspended in 400 ml of infiltration medium (2.2 g Murashige and Skoog medium, 50 g sucrose, 50 μl Silwet copolymer L-77, 10 μl of 1 mg/ml BAP and 0.5 g MES per litre). Flowering A. thaliana plants were immersed for 1 minute into the suspension by inverting the pot and then left standing, covered with a plastic bag, to maintain the humidity. Next day the plastic bag was removed. This was done with 5 pots per construct. Two ecotypes, c-24 and Col-0 were transformed with the amplicon constructs this way.

REFERENCES

[0148] Inasmuch as they may be required by the person skilled in the art to practice the present invention, all citations are specifically included herein by cross-reference.

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[0154] Frisch, D. A., L. W. Harris-Haller, et al. (1995). “Complete Sequence of the binary vector Bin 19.” Plant Molecular Biology 27: 405-409.

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[0160] Kjemtrup, S., Sampson, K. S., Peele, C. G., Nguyen, L. V., Conkling, M. A., Thompson, W. F., and Robertson, D. (1998). Gene silencing from plant DNA carried by a geminivirus. Plant J. 14, 91-100.

[0161] Kumagai, M. H., Donson, J., Della-Cioppa, G., Harvey, D., Hanley, K., and Grill, L. K. (1995). Cytoplasmic inhibition of carotenoid biosynthesis with virus-derived RNA. Proc.Natl.Acad.Sci.USA 92, 1679-1683.

[0162] Mori, M., Zhang, G.-H., Kaido, M., Okuno, T., and Furusawa, I. (1993). Efficient production of human gamma interferon in tobacco protoplasts by genetically engineered brome mosaic virus RNAs. J.Gen.Virol. 74, 1255-1260.

[0163] Napoli, C., Lemieux, C., and Jorgensen, R. A. (1990). Introduction of a chimeric chalcone synthase gene into Petunia results in reversible co-suppression of homologous genes in trans. Plant Cell 2, 279-289.

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[0168] Sequence Appendix Key to the sequence annotation: Lower-case plasmid backbone sequence. Lower-case italics sequence inserted into the amplicon constructs. Lower-case underlined CaMV 35S promoter sequence. UPPER-CASE tobacco rattle virus RNA1 cDNA sequence. UPPER-CASE AND BOLD Arabidopsis NIA1-intron 3 sequence. UPPER-CASE ITALICS UNDERLINED multiple cloning site. Lower-case and bold CaMV 35S terminator sequence. . . . deletion oE 16K. (. . . ) deletion of the replicase and intron.

[0169] The sequence of pBTAΔMP is given in full, including the vector backbone. For the sequence of the other three amplicon constructs (pBTAΔMPΔ16K, pBTAΔRepΔMP and pBTAΔRepΔMPΔ16K) vector backbone is not given, since is the same for all of them. SEQ ID NO 1 - Complete Sequence of pBINTRA6 Plasmid. tactccaaaaatgtcaaagatacagtctcagaagaccaaagggctattgagacttttcaacaaaggg taatttcgggaaacctcctcggattccattgcccagctatctgtcacttcatcgaaaggacagtaga aaaggaaggtggctcctacaaatgccatcattgcgataaaggaaaggctatcattcaagatgcctct gccgacagtggtcccaaagatggacccccacccacgaggagcatcgtggaaaaagaagacgtcccaa ccacgtcttcaaagcaagtggattgatgtgacatctccactgacgtaagggatgacgcacaatccca ctatccttcgcaagacccttcttctatataaggaagttcatttcatttggagaggacagcccaagct ttctagagGATCCATAAAACATTTCAATCCTTTGAACGCGGTAGAACGTGCTAATTGGATTTTGGTG AGAACGCGGTAGAACGTACTTATCACCTACAGTTTTATTTTGTTTTTCTTTTTGGTTTAATCTATCC AGCTTAGTACCGAGTGGGGGAAAGTGACTGGTGTGCCTAAAACCTTTTCTTTGATACTTTGTAAAAA TACATACAGATACAATGGCGAACGGTAACTTCAAGTTGTCTCAATTGCTCAATGTGGACGAGATGTC TGCTGAGCAGAGGAGTCATTTCTTTGACTTGATGCTGACTAAACCTGATTGTGAGATCGGGCAAATG ATGCAAAGAGTTGTTGTTGATAAAGTCGATGACATGATTAGAGAAAGAAAGACTAAAGATCCAGTGA TTGTTCATGAAGTTCTTTCTCAGAAGGAACAGAACAAGTTGATGGAAATTTATCCTGAATTCAATAT CGTGTTTAAAGACGACAAAAACATGGTTCATGGGTTTGCGGCTGCTGAGCGAAAACTACAAGCTTTA TTGCTTTTAGATAGAGTTCCTGCTCTGCAAGAGGTGGATGACATCGGTGGTCAATGGTCGTTTTGGG TAACTAGAGGTGAGAAAAGGATTCATTCCTGTTGTCCAAATCTAGATATTCGGGATGATCAGAGAGA AATTTCTCGACAGATATTTCTTACTGCTATTGGTGATCAAGCTAGAAGTGGTAAGAGACAGATGTCG GAGAATGAGCTGTGGATGTATGACCAATTTCGTGAAAATATTGCTGCGCCTAACGCGGTTAGGTGCA ATAATACATATCAGGGTTGTACATGTAGGGGTTTTTCTGATGGTAAGAAGAAAGGCGCGCAGTATGC GATAGCTCTTCACAGCCTGTATGACTTCAAGTTGAAAGACTTGATGGCTACTATGGTTGAGAAGAAA ACTAAAGTGGTTCATGCTGCTATGCTTTTTGCTCCTGAAAGTATGTTAGTGGACGAAGGTCCATTAC CTTCTGTTGACGGTTACTACATGAAGAAGAACGGGAAGATCTATTTCGGTTTTGAGAAAGATCCTTC CTTTTCTTACATTCATGACTGGGAAGAGTACAAGAAGTATCTACTGGGGAAGCCAGTGAGTTACCAA GGGAATGTGTTCTACTTCGAACCGTGGCAGGTGAGAGGAGACACAATGCTTTTTTCGATCTACAGGA TAGCTGGAGTTCCGAGGAGGTCTCTATCATCGCAAGAGTACTACCGAAGAATATATATCAGTAGATG GGAAAGCATGGTTGTTGTCCCAATTTTCGATCTGGTCGAATCAACGCGAGAGTTGGTCAAGAAAGAC CTGTTTGTAGAGAAACAATTCATGGACAAGTGTTTGGATTACATAGCTAGGTTATCTGACCAGCAGC TGACCATAAGCAATGTTAAATCATACTTGAGTTCAAATAATTGGGTCTTATTCATAAACGGGGCGGC CGTGAAGAACAAGCAAAGTGTAGATTCTCGAGATTTACAGTTGTTGGCTCAAACTTTGCTAGTGAAG GAACAAGTGGCGAGACCTGTCATGAGGGAGTTGCGTGAAGCAATTCTGACTGAGACGAAACCTATCA CGTCATTGACTGATGTGCTGGGTTTAATATCAAGAAAACTGTGGAAGCAGTTTGCTAACAAGATCGC AGTCGGCGGATTCGTTGGCATGGTTGGTACTCTAATTGGATTCTATCCAAAGAAGGTACTAACCTGG GCGAAGGACACACCAAATGGTCCAGAACTATGTTACGAGAACTCGCACAAAACCAAGGTGATAGTAT TTCTGAGTGTTGTGTATGCCATTGGAGGAATCACGCTTATGCGTCGAGACATCCGAGATGGACTGGT GAAAAAACTATGTGATATGTTTGATATCAAACGGGGGGCCCATGTCTTAGACGTTGAGAATCCGTGC CGCTATTATGAAATCAACGATTTCTTTAGCAGTCTGTATTCGGCATCTGAGTCCGGTGAGACCGTTT TACCAGATTTATCCGAGGTAAAAGCCAAGTCTGATAAGCTATTGCAGCAGAAGAAAGAAATCGCTGA CGAGTTTCTAAGTGCAAAATTCTCTAACTATTCTGGCAGTTCGGTGAGAACTTCTCCACCATCGGTG GTCGGTTCATCTCGAAGCGGACTGGGTCTGTTGTTGGAAGACAGTAACGTGCTGACCCAAGCTAGAG TTGGAGTTTCAAGAAAGGTAGACGATGAGGAGATCATGGAGCAGTTTCTGAGTGGTCTTATTGACAC TGAAGCAGAAATTGACGAGGTTGTTTCAGCCTTTTCAGCTGAATGTGAAAGAGGGGAAACAAGCGGT ACAAAGGTGTTGTGTAAACCTTTAACGCCACCAGGATTTGAGAACGTGTTGCCAGCTGTCAAACCTT TGGTCAGCAAAGGAAAAACGGTCAAACGTGTCGATTACTTCCAAGTGATGGGAGGTGAGAGATTACC AAAAAGGCCGGTTGTCAGTGGAGACGATTCTGTGGACGCTAGAAGAGAGTTTCTGTACTACTTAGAT GCGGAGAGAGTCGCTCAAAATGATGAAATTATGTCTCTGTATCGTGACTATTCGAGAGGAGTTATTC GAACTGGAGGTCAGAATTACCCGCACGGACTGGGAGTGTGGGATGTGGAGATGAAGAACTGGTGCAT ACGTCCAGTGGTCACTGAACATGCTTATGTGTTCCAACCAGACAAACGTATGGATGATTGGTCGGGA TACTTAGAAGTGGCTGTTTGGGAACGAGGTATGTTGGTCAACGACTTCGCGGTCGAAAGGATGAGTG ATTATGTCATAGTTTGCGATCAGACGTATCTTTGCAATAACAGGTAATAATCCTCTCTCTTGATATT TTTAAATTATAGAATTAATTAGTTTACTTTATTCTTTACTATATGATTTAAATAGTTTAATCTTGTT TTTGAGTAAACTATTCGATTTTGATATTTGTATTCGTCCTACAAAGTTGGAAATACTGATGATATTT TCTTTTGAACGTGATACCTACCAATACTAATCTTACGGAATCTTTTAATAGAGCACTAATCAACATG GAACTAAAGACCAATTCTTAAGTGTCTCTGTTGTACAGTTCATTTTAGTAGTGCGTTTAAGTATTAT TATCTCCCTTCATGCGGGGCAATTATGTAGATTAAAATCGAAATTATATAAAATTTACATAAGTCTA AGTCTAGGGTCTCCAGCTAATTGTTATTTTTTTAACGATGTTGACTAAAGCAATAACGACGTTGACT TGTGTTAAACAGGTTGATCTTGGACAATTTAAGTGCCCTGGATCTAGGACCAGTTAACTGTTCTTTT GAATTAGTTGACGGTGTACCTGGTTGTGGTAAGTCGACAATGATTGTCAACTCAGCTAATCCTTGTG TCGATGTGGTTCTCTCTACTGGGAGAGCAGCAACCGACGACTTGATCGAGAGATTCGCGAGCAAAGG TTTTCCATGCAAATTGAAAAGGAGAGTGAAGACGGTTGATTCTTTTTTGATGCATTGTGTCGATGGT TCTTTAACCGGAGACGTGTTGCATTTCGACGAAGCTCTCATGGCCCATGCTGGTATGGTGTACTTTT GCGCTCAGATAGCTGGTGCTAAACGATGTATCTGTCAAGGAGATCAGAATCAAATTTCTTTCAAGCC TAGGGTATCTCAAGTTGATTTGAGGTTTTCTAGTCTGGTCGGAAAGTTTGACATTGTTACAGAAAAA AGAGAAACTTACAGAAGTCCAGCAGATGTGGCTGCCGTATTGAACAAGTACTATACTGGAGATGTCA GAACACATAACGCGACTGCTAATTCGATGACGGTGAGGAAGATTGTGTCTAAAGAACAGGTTTCTTT GAAGCCTGGTGCTCAGTACATAACTTTCCTTCAGTCTGAGAAGAAGGAGTTGGTAAATTTGTTGGCA TTGAGGAAAGTGGCAGCTAAAGTGAGTACAGTACACGAGTCGCAAGGAGAGACATTCAAAGATGTAG TCCTAGTCAGGACGAAACCTACGGATGACTCAATCGCTAGAGGTCGGGAGTACTTAATCGTGGCATT GTCGCGTCACACACAATCACTTGTGTATGAAACTGTGAAAGAGGACGATGTAAGCAAAGAGATCAGG GAAAGTGCCGCGCTTACGAAGGCGGCTTTGGCAAGATTTTTTGTTACTGAGACCGTCTTATGACGGT TTCGGTCTAGGTTTGATGTCTTTAGACATCATGAAGGGCCTTGCGCCGTTCCAGATTCAGGTACGAT TACGGACTTGGAGATGTGGTACGACGCTTTGTTTCCGGGAAATTCGTTAAGAGACTCAAGCCTAGAC GGGTATTTGGTGGCAACGACTGATTGCAATTTGCGATTAGACAATGTTACGATCAAAAGTGGAAACT GGAAAGACAAGTTTGCTGAAAAAGAAACGTTTCTGAAACCGGTTATTCGTACTGCTATGCCTGACAA AAGGAAGACTACTCAGTTGGAGAGTTTGTTAGCATTGCAGAAAAGGAACCAAGCGGCACCCGATCTA CAAGAAAATGTGCACGCGACAGTTCTAATCGAAGAGACGATGAAGAAGCTGAAATCTGTTGTCTACG ATGTGGGAAAAATTCGGGCTGATCCTATTGTCAATAGAGCTCAAATGGAGAGATGGTGGAGAAATCA AAGCACAGCGGTACAGGCTAAGGTAGTAGCAGATGTGAGAGAGTTACATGAAATAGACTATTCGTCT TACATGTATATGATCAAATCTGACGTGAAACCTAAGACTGATTTAACACCGCAATTTGAATACTCAG CTCTACAGACTGTTGTGTATCACGAGAAGTTGATCAACTCGTTGTTCGGTCCAATTTTCAAAGAAAT TAATGAACGCAAGTTGGATGCTATGCAACCACATTTTGTGTTCAACACGAGAATGACATCGAGTGAT TTAAACGATCGAGTGAAGTTCTTAAATACGGAAGCGGCTTACGACTTTGTTGAGATAGACATGTCTA AATTCGACAAGTCGGCAAATCGCTTCCATTTACAACTGCAGCTGGAGATTTACAGGTTATTTGGGCT GGATGAGTGGGCGGCCTTCCTTTGGGAGGTGTCGCACACTCAAACTACTGTGAGAGATATTCAAAAT GGTATGATGGCGCATATTTGGTACCAACAAAAGAGTGGAGATGCTGATACTTATAATGCAAATTCAG ATAGAACACTGTGTGCACTCTTGTCTGAATTACCATTGGAGAAAGCAGTCATGGTTACATATGGAGG AGATGACTCACTGATTGCGTTTCCTAGAGGAACGCAGTTTGTTGATCCGTGTCCAAAGTTGGCTACT AAGTGGAATTTCGAGTGCAAGATTTTTAAGTACGATGTCCCAATGTTTTGTGGGAAGTTCTTGCTTA AGACGTCATCGTGTTACGAGTTCGTGCCAGATCCGGTAAAAGTTCTGACGAAGTTGGGGAAAAAGAG TATAAAGGATGTGCAACATTTAGCCGAGATCTACATCTCGCTGAATGATTCCAATAGAGCTCTTGGG AACTACATGGTGGTATCCAAACTGTCCGAGTCTGTTTCAGACCGGTATTTGTACAAAGGTGATTCTG TTCATGCGCTTTGTGCGCTATGGAAGCATATTAAGAGTTTTACAGCTCTGTGTACATTATTCCGAGA CGAAAACGATAAGGAATTGAACCCGGCTAAGGTTGATTGGAAGAAGGCACAGAGAGCTGTGTCAAAC TTTTACGACTGGTAATATGGAAGACAAGTCATTGGTCACCTTGAAGAAGAAGACTTTCGAAGTCTCA AAATTCTCAAATCTAGGGGCCATTGAATTGTTTGTGGACGGTAGGAGGAAGAGACCGAAGTATTTTC ACAGAAGAAGAGAAACTGTCCTAAATCATGTTGGTGGGAAGAAGAGTGAACACAAGTTAGACGTTTT TGACCAAAGGGATTACAAAATGATTAAATCTTACGCGTTTCTAAAGGTAGTAGGTGTACAACTAGTT GTAACATCACATCTACCTGCAGATACGCCTGGGTTCATTCAAATCGATCTGTTGGATTCGAGACTTA CTGAGAAAAGAAAGAGAGGAAAGACTATTCAGAGATTCAAAGCTCGAGCTTGCGATAACTGTTCAGT TGCGCAGTACAAGGTTGAATACAGTATTTCCACACAGGAGAACGTACTTGATGTCTGGAAGGTGGGT TGTATTTCTGAGGGCGTTCCGGTCTGTGACGGTACATACCCTTTCAGTATCGAAGTGTCGCTAATAT GGGTTGCTACTGATTCGACTAGGCGCCTCAATGTGGAAGAACTGAACAGTTCGGATTACATTGAAGG CGATTTTACCGATCAAGAGGTTTTCGGTGAGTTCATGTCTTTGAAACAAGTGGAGATGAAGACGATT GAGGCGAAGTACGATGGTCCTTACAGACCAGCTACTACTAGACCTAAGTCATTATTGTCAAGTGAAG ATGTTAAGAGAGCGTCTAATAAGAAAAACTCGTCTTAATGCATAAAGAAATTTATTGTCAATATGAC GTGTGTACTCAAGGGTTGTGTGAATGAAGTCACTGTTCTTGGTCACGAGACGTGTAGTATCGGTCAT GCTAACAAATTGCGAAAGCAAGTTGCTGACATGGTTGGTGTCACACGTAGGTGTGCGGAAAATAATT GTGGATGGTTTGTCTGTGTTGTTATCAATGATTTTACTTTTGATGTGTATAATTGTTGTGGCCGTAG TCACCTTGAAAAGTGTCGTAAACGTGTTGAAACAAGAAATCGAGAAATTTGGAAACAAATTCGACGA AATCAAGCTGAAAACATGTCTGCGACAGCTAAAAAGTCTCATAATTCGAAGACCTCTAAGAAGAAAT TCAAAGAGGACAGAGAATTTGGGACACCAAAAAGATTTTTAAGAGATGATGTTCCTTTCGGGATTGA TCGTTTGTTTGCTTTTTGATTTTATTTTATATTGTTATCTGTTTCTGTGTATAGACTGTTTGAGATT GGCGCTTGGCCGACTCATTGTCTTACCATAGGGGAACGGACTTTGTTTGTGTTGTTATTTTATTTGT ATTTTATTAAAATTCTCAATGATCTGAAAAGGCCTCGAGGCTAAGAGATTATTGGCGGGTGAGTAAG TACTTTTAAAGTGATGATGGTTACAAAGGCAAAAGGGGTAAAACCCCTCGCCTACGTAAGCGTTATT ACGCCCGgatcccccggggagctcgaattcgctgaaatcaccagtctctctctacaaatctatctct ctctattttttccataaataatgtgtgagtagtttcccgataagggaaattagggttcttatagggt ttcgetcatgtgttgagcatataagaaacccttagtatgtatttgtatttgtaaaatacttctatta tcaataaaatttctaattcctaaaaccaaaatccagtactaaaatccagatctcctaaagtccctat agatctttgtcgtgaatataaaccagacacgagacgactaaacdtggagcccagacgccgttcgaag ctagaagtaccgcttaggcaggaggccgttagggaaaagatgctaaggcagggttggttacgttgac tcccccgtaggtttggtttaaatatgatgaagtggacggaaggaaggaggaagacaaggaaggataa ggttgcaggccctgtgcaaggtaagaagatggaaatttgatagaggtacgctactatacttatacta tacgctaagggaatgcttgtatttataccctataccccctaataaccccttatcaatttaagaaata atccgcataagcccccgcttaaaaattggtatcagagccatgaataggtctatgaccaaaactcaag aggataaaacctcaccaaaatacgaaagagttcttaactctaaagataaaagatctttcaagatcaa aactagttccctcacaccggagcatgcgatatcctcgacctgcaggcatgcaagcttggcgtaatca tggtcatagctgtttcctgtgtgaaattgttatccgctcacaattccacacaacatacgagccggaa gcataaagtgtaaagcctggggtgcctaatgagtgagctaactcacattaattgcgttgcgctcact gcccgctttccagtcgggaaacctgtcgtgccagctgcattaatgaatcggccaacgcgcggggaga ggcggtttgcgtattgggccaaagacaaaagggcgacattcaaccgattgagggagggaaggtaaat attgacggaaattattcattaaaggtgaattatcaccgtcaccgacttgagccatttgggaattaga gccagcaaaatcaccagtagcaccattaccattagcaaggccggaaacgtcaccaatgaaaccatcg atagcagcaccgtaatcagtagcgacagaatcaagtttgcctttagcgtcagactgtagcgcgtttt catcggcattttcggtcatagcccccttattagcgtttgccatcttttcataatcaaaatcaccgga accagagccaccaccggaaccgcctccctcagagccgccaccctcagaaccgccaccctcagagcca ccaccctcagagccgccaccagaaccaccaccagagccgccgccagcattgacaggaggcccgatct agtaacatagatgacaccgcgcgcgataatttatcctagtttgcgcgctatattttgttttctatcg cgtattaaatgtataattgcgggactctaatcataaaaacccatctcataaataacgtcatgcatta catgttaattattacatgcttaacgtaattcaacagaaattatatgataatcatcgcaagaccggca acaggattcaatcttaagaaactttattgccaaatgtttgaacgatcggggatcatccgggtctgtg gcgggaactccacgaaaatatccgaacgcagcaagatatcgcggtgcatctcggtcttgcctgggca gtcgccgccgacgccgttgatgtggacgccgggcccgatcatattgtcgctcaggatcgtggcgttg tgcttgtcggccgttgctgtcgtaatgatatcggcaccttcgaccgcctgttccgcagagatcccgt gggcgaagaactccagcatgagatccccgcgctggaggatcatccagccggcgtcccggaaaacgat tccgaagcccaacctttcatagaaggcggcggtggaatcgaaatctcgtgatggcaggttgggcgtc gcttggtcggtcatttcgaaccccagagtcccgctcagaagaactcgtcaagaaggcgatagaaggc gatgcgctgcgaatcgggagcggcgataccgtaaagcacgaggaagcggtcagcccattcgccgcca agctcttcagcaatatcacgggtagccaacgctatgtcctgatagcggtccgccacacccagccggc cacagtcgatgaatccagaaaagcggccattttccaccatgatattcggcaagcaggcatcgccatg ggtcacgacgagatcatcgccgtcgggcatgcgcgccttgagcctggcgaacagttcggctggcgcg agcccctgatgctcttcgtccagatcatcctgatcgacaagaccggcttccatccgagtacgtgctc gctcgatgcgatgtttcgcttggtggtcgaatgggcaggtagccggatcaagcgtatgcagccgccg cattgcatcagccatgatggatactttctcggcaggagcaaggtgagatgacaggagatcctgcccc ggcacttcgcccaatagcagccagtcccttcccgcttcagtgacaacgtcgagcacagctgcgcaag gaacgcccgtcgtggccagccacgatagccgcgctgcctcgtcctgcagttcattcagggcaccgga caggtcggtcttgacaaaaagaaccgggcgcccctgcgctgacagccggaacacggcggcatcagag cagccgattgtctgttgtgcccagtcatagccgaatagcctctccacccaagcggccggagaacctg cgtgcaatccatcttgttcaatcatgcgaaacgatccagatccggtgcagattatttggattgagag tgaatatgagactctaattggataccgaggggaatttatggaacgtcagtggagcatttttgacaag aaatatttgctagctgatagtgaccttaggcgacttttgaacgcgcaataatggtttctgacgtatg tgcttagctcattaaactccagaaacccgcggctgagtggctccttcaacgttgcggttctgtcagt tccaaacgtaaaacggcttgtcccgcgtcatcggcgggggtcataacgtgactcccttaattctccg ctcatgatcagattgtcgtttcccgccttcagtttaaactatcagtgtttgacaggatatattggcg ggtaaacctaagagaaaagagcgtttattagaataatcggatatttaaaagggcgtgaaaaggttta tccgttcgtccatttgtatgtgcatgccaaccacagggttccccagatctggcgccggccagcgaga cgagcaagattggccgccgcccgaaacgatccgacagcgcgcccagcacaggtgcgcaggcaaattg caccaacgcatacagcgccagcagaatgccatagtgggcggtgacgtcgttcgagtgaaccagatcg cgcaggaggcccggcagcaccggcataatcaggccgatgccgacagcgtcgagcgcgacagtgctca gaattacgatcaggggtatgttgggtttcacgtctggcctccggaccagcctccgctggtccgattg aacgcgcggattctttatcactgataagttggtggacatattatgtttatcagtgataaagtgtcaa gcatgacaaagttgcagccgaatacagtgatccgtgccgccctggacctgttgaacgaggtcggcgt agacggtctgacgacacgcaaactggcggaacggttgggggttcagcagccggcgctttactggcac ttcaggaacaagcgggcgctgctcgacgcactggccgaagccatgctggcggagaatcatacgcatt cggtgccgagagccgacgacgactggcgctcatttctgatcgggaatgcccgcagcttcaggcaggc gctgctcgcctaccgcgatggcgcgcgcatccatgccggcacgcgaccgggcgcaccgcagatggaa acggccgacgcgcagcttcgcttcctctgcgaggcgggtttttcggccggggacgccgtcaatgcgc tgatgacaatcagctacttcactgttggggccgtgcttgaggagcaggccggcgacagcgatgccgg cgagcgcggcggcaccgttgaacaggctccgctctcgccgctgttgcgggccgcgatagacgccttc gacgaagccggtccggacgcagcgttcgagcagggactcgcggtgattgtcgatggattggcgaaaa ggaggctcgttgtcaggaacgttgaaggaccgagaaagggtgacgattgatcaggaccgctgccgga gcgcaacccactcactacagcagagccatgtagacaacatcccctccccctttccaccgcgtcagac gcccgtagcagcccgctacgggctttttcatgccctgccctagcgtccaagcctcacggccgcgctc ggcctctctggcggccttctggcgctcttccgcttcctcgctcactgactcgctgcgctcggtcgtt cggctgcggcgagcggtatcagctcactcaaaggcggtaatacggttatccacagaatcaggggata acgcaggaaagaacatgtgagcaaaaggccagcaaaaggccaggaaccgtaaaaaggccgcgttgct ggcgtttttccataggctccgcccccctgacgagcatcacaaaaatcgacgctcaagtcagaggtgg cgaaacccgacaggactataaagataccaggcgtttccccctggaagctccctcgtgcgctctcctg ttccgaccctgccgcttaccggatacctgtccgcctttctcccttcgggaagcgtggcgcttttccg ctgcataaccctgcttcggggtcattatagcgattttttcggtatatccatcctttttcgcacgata tacaggattttgccaaagggttcgtgtagactttccttggtgtatccaacggcgtcagccgggcagg ataggtgaagtaggcccacccgcgagcgggtgttccttcttcactgtcccttattcgcacctggcgg tgctcaacgggaatcctgctctgcgaggctggccggctaccgccggcgtaacagatgagggcaagcg gatggctgatgaaaccaagccaaccaggaagggcagcccacctatcaaggtgtactgccttccagac gaacgaagagcgattgaggaaaaggcggcggcggccggcatgagcctgtcggcctacctgctggccg tcggccagggctacaaaatcacgggcgtcgtggactatgagcacgtccgcgagctggcccgcatcaa tggcgacctgggccgcctgggcggcctgctgaaactctggctcaccgacgacccgcgcacggcgcgg ttcggtgatgccacgatcctcgccctgctggcgaagatcgaagagaagcaggacgagcttggcaagg tcatgatgggcgtggtccgcccgagggcagagccatgacttttttagccgctaaaacggccgggggg tgcgcgtgattgccaagcacgtccccatgcgctccatcaagaagagcgacttcgcggagctggtgaa gtacatcaccgacgagcaaggcaagaccgagcgcctttgcgacgctcaccgggctggttgccctcgc cgctgggctggcggccgtctatggccctgcaaacgcgccagaaacgccgtcgaagccgtgtgcgaga caccgcggccgccggcgttgtggatacctcgcggaaaacttggccctcactgacagatgaggggcgg acgttgacacttgaggggccgactcacccggcgcggcgttgacagatgaggggcaggctcgatttcg gccggcgacgtggagctggccagcctcgcaaatcggcgaaaacgcctgattttacgcgagtttccca cagatgatgtggacaagcctggggataagtgccctgcggtattgacacttgaggggcgcgactactg acagatgaggggcgcgatccttgacacttgaggggcagagtgctgacagatgaggggcgcacctatt gacatttgaggggctgtccacaggcagaaaatccagcatttgcaagggtttccgcccgtttttcggc caccgctaacctgtcttttaacctgcttttaaaccaatatttataaaccttgtttttaaccagggct gcgccctgtgcgcgtgaccgcgcacgccgaaggggggtgcccccccttctcgaaccctcccggcccg ctaacgcgggcctcccatccccccaggggctgcgcccctcggccgcgaacggcctcaccccaaaaat ggcagcgctggcagtccttgccattgccgggatcggggcagtaacgggatgggcgatcagcccgagc gcgacgcccggaagcattgacgtgccgcaggtgctggcatcgacattcagcgaccaggtgccgggca gtgagggcggcggcctgggtggcggcctgcccttcacttcggccgtcggggcattcacggacttcat ggcggggccggcaatttttaccttgggcattcttggcatagtggtcgcgggtgccgtgctcgtgttc gggggtgcgataaacccagcgaaccatttgaggtgataggtaagattataccgaggtatgaaaacga gaattggacctttacagaattactctatgaagcgccatatttaaaaagctaccaagacgaagaggat gaagaggatgaggaggcagattgccttgaatatattgacaatactgataagataatatatcttttat atagaagatatcgccgtatgtaaggatttcagggggcaaggcataggcagcgcgcttatcaatatat ctatagaatgggcaaagcataaaaacttgcatggactaatgcttgaaacccaggacaataaccttat agcttgtaaattctatcataattgggtaatgactccaacttattgatagtgttttatgttcagataa tgcccgatgactttgtcatgcagctccaccgattttgagaacgacagcgacttccgtcccagccgtg ccaggtgctgcctcagattcaggttatgccgctcaattcgctgcgtatatcgcttgctgattacgtg cagctttcccttcaggcgggattcatacagcggccagccatccgtcatccatatcaccacgtcaaag ggtgacagcaggctcataagacgccccagcgtcgccatagtgcgttcaccgaatacgtgcgcaacaa ccgtcttccggagactgtcatacgcgtaaaacagccagcgctggcgcgatttagccccgacatagcc ccactgttcgtccatttccgcgcagacgatgacgtcactgcccggctgtatgcgcgaggttaccgac tgcggcctgagttttttaagtgacgtaaaatcgtgttgaggccaacgcccataatgcgggctgttgc ccggcatccaacgccattcatggccatatcaatgattttctggtgcgtaccgggttgagaagcggtg taagtgaactgcagttgccatgttttacggcagtgagagcagagatagcgctgatgtccggcggtgc ttttgccgttacgcaccaccccgtcagtagctgaacaggagggacagctgatagacacagaagccac tggagcacctcaaaaacaccatcatacactaaatcagtaagttggcagcatcacccataattgtggt ttcaaaatcggctccgtcgatactatgttatacgccaactttgaaaacaactttgaaaaagctgttt tctggtatttaaggttttagaatgcaaggaacagtgaattggagttcgtcttgttataattagcttc ttggggtatctttaaatactgtagaaaagaggaaggaaataataaatggctaaaatgagaatatcac cggaattgaaaaaactgatcgaaaaataccgctgcgtaaaagatacggaaggaatgtctcctgctaa ggtatataagctggtgggagaaaatgaaaacctatatttaaaaatgacggacagccggtataaaggg accacctatgatgtggaacgggaaaaggacatgatgctatggctggaaggaaagctgcctgttccaa aggtcctgcactttgaacggcatgatggctggagcaatctgctcatgagtgaggccgatggcgtcct ttgctcggaagagtatgaagatgaacaaagccctgaaaagattatcgagctgtatgcggagtgcatc aggctctttcactccatcgacatatcggattgtccctatacgaatagcttagacagccgcttagccg aattggattacttactgaataacgatctggccgatgtggattgcgaaaactgggaagaagacactcc atttaaagatccgcgcgagctgtatgattttttaaagacggaaaagcccgaagaggaacttgtcttt tcccacggcgacctgggagacagcaacatctttgtgaaagatggcaaagtaagtggctttattgatc ttgggagaagcggcagggcggacaagtggtatgacattgccttctgcgtccggtcgatcagggagga tatcggggaagaacagtatgtcgagctattttttgacttactggggatcaagcctgattgggagaaa ataaaatattatattttactggatgaattgttttagtacctagatgtggcgcaacgatgccggcgac aagcaggagcgcaccgacttcttccgcatcaagtgttttggctctcaggccgaggcccacggcaagt atttgggcaaggggtcgctggtattcgtgcagggcaagattcggaataccaagtacgagaaggacgg ccagacggtctacgggaccgacttcattgccgataaggtggattatctggacaccaaggcaccaggc gggtcaaatcaggaataagggcacattgccccggcgtgagtcggggcaatcccgcaaggagggtgaa tgaatcggacgtttgaccggaaggcatacaggcaagaactgatcgacgcggggttttccgccgagga tgccgaaaccatcgcaagccgcaccgtcatgcgtgcgccccgcgaaaccttccagtccgtcggctcg atggtccagcaagctacggccaagatcgagcgcgacagcgtgcaactggctccccctgccctgcccg cgccatcggccgccgtggagcgttcgcgtcgtctcgaacaggaggcggcaggtttggcgaagtcgat gaccatcgacacgcgaggaactatgacgaccaagaagcgaaaaaccgccggcgaggacctggcaaaa caggtcagcgaggccaagcaggccgcgttgctgaaacacacgaagcagcagatcaaggaaatgcagc tttccttgttcgatattgcgccgtggccggacacgatgcgagcgatgccaaacgacacggcccgctc tgccctgttcaccacgcgcaacaagaaaatcccgcgcgaggcgctgcaaaacaaggtcattttccac gtcaacaaggacgtgaagatcacctacaccggcgtcgagctgcgggccgacgatgacgaactggtgt ggcagcaggtgttggagtacgcgaagcgcacccctatcggcgagccgatcaccttcacgttctacga gctttgccaggacctgggctggtcgatcaatggccggtattacacgaaggccgaggaatgcctgtcg cgcctacaggcgacggcgatgggcttcacgtccgaccgcgttgggcacctggaatcggtgtcgctgc tgcaccgcttccgcgtcctggaccgtggcaagaaaacgtcccgttgccaggtcctgatcgacgagga aatcgtcgtgctgtttgctggcgaccactacacgaaattcatatgggagaagtaccgcaagctgtcg ccgacggcccgacggatgttcgactatttcagctcgcaccgggagccgtacccgctcaagctggaaa ccttccgcctcatgtgcggatcggattccacccgcgtgaagaagtggcgcgagcaggtcggcgaagc ctgcgaagagttgcgaggcagcggcctggtggaacacgcctgggtcaatgatgacctggtgcattgc aaacgctagggccttgtggggtcagttccggctgggggttcagcagccagcgctttactggcatttc aggaacaagcgggcactgctcgacgcacttgcttcgctcagtatcgctcgggacgcacggcgcgctc tacgaactgccgataaacagaggattaaaattgacaattgtgattaaggctcagattcgacggcttg gagcggccgacgtgcaggatttccgcgagatccgattgtcggccctgaagaaagctccagagatgtt cgggtccgtttacgagcacgaggagaaaaagcccatggaggcgttcgctgaacggttgcgagatgcc gtggcattcggcgcctacatcgacggcgagatcattgggctgtcggtcttcaaacaggaggacggcc ccaaggacgctcacaaggcgcatctgtccggcgttttcgtggagcccgaacagcgaggccgaggggt cgccggtatgctgctgcgggcgttgccggcgggtttattgctcgtgatgatcgtccgacagattcca acgggaatctggtggatgcgcatcttcatcctcggcgcacttaatatttcgctattctggagcttgt tgtttatttcggtctaccgcctgccgggcggggtcgcggcgacggtaggcgctgtgcagccgctgat ggtcgtgttcatctctgccgctctgctaggtagcccgatacgattgatggcggtcctgggggctatt tgcggaactgcgggcgtggcgctgttggtgttgacaccaaacgcagcgctagatcctgtcggcgtcg cagcgggcctggcgggggcggtttccatggcgttcggaaccgtgctgacccgcaagtggcaacctcc cgtgcctctgctcacctttaccgcctggcaactggcggccggaggacttctgctcgttccagtagct ttagtgtttgatccgccaatcccgatgcctacaggaaccaatgttctcggcctggcgtggctcggcc tgatcggagcgggtttaacctacttcctttggttccgggggatctcgcgactcgaacctacagttgt ttccttactgggctttctcagccccagatctggggtcgatcagccggggatgcatcaggccgacagt cggaacttcgggtccccgacctgtaccattcggtgagcaatggataggggagttgatatcgtcaacg ttcacttctaaagaaatagcgccactcagcttcctcagcggctttatccagcgatttcctattatgt cggcatagttctcaagatcgacagcctgtcacggttaagcgagaaatgaataagaaggctgataatt cggatctctgcgagggagatgatatttgatcacaggcagcaacgctctgtcatcgttacaatcaaca tgctaccctccgcgagatcatccgtgtttcaaacccggcagcttagttgccgttcttccgaatagca tcggtaacatgagcaaagtctgccgccttacaacggctctcccgctgacgccgtcccggactgatgg gctgcctgtatcgagtggtgattttgtgccgagctgccggtcggggagctgttggctggctggtggc aggatatattgtggtgtaaacaaattgacgcttagacaacttaataacacattgcggacgtttttaa tgtactggggtggtttttcttttcaccagtgagacgggcaacagctgattgcccttcaccgcctggc cctgagagagttgcagcaagcggtccacgctggtttgccccagcaggcgaaaatcctgtttgatggt ggttccgaaatcggcaaaatcccttataaatcaaaagaatagcccgagatagggttgagtgttgttc cagtttggaacaagagtccactattaaagaacgtggactccaacgtcaaagggcgaaaaaccgtcta tcagggcgatggcccactacgtgaaccatcacccaaatcaagttttttggggtcgaggtgccgtaaa gcactaaatcggaaccctaaagggagcccccgatttagagcttgacggggaaagccggcgaacgtgg cgagaaaggaagggaagaaagcgaaaggagcgggcgccattcaggctgcgcaactgttgggaagggc gatcggtgcgggcctcttcgctattacgccagctggcgaaagggggatgtgctgcaaggcgattaag ttgggtaacgccagggttttcccagtcacgacgttgtaaaacgacggccagtgaattcgagctcggt acccccc

Amplicon Constructs

[0170] SEQ ID NO 2 - pBTAΔMP tactccaaaaatgtcaaagatacagtctcagaagaccaaagggctattgagacttttcaacaaaggg taatttcgggaaacctcctcggattccattgcccagctatctgtcacttcatcgaaaggacagtaga aaaggaaggtggctcctacaaatgccatcattgcgataaaggaaaggctatcattcaagatgcctct gccgacagtggtcccaaagatggacccccacccacgaggagcatcgtggaaaaagaagacgtcccaa ccacgtcttcaaagcaagtggattgatgtgacatctccactgacgtaagggatgacgcacaatccca ctatccttcgcaagacccttcttctatataaggaagttcatttcatttggagaggacagcccaagct ttctagagGATCCATAAAACATTTCAATCCTTTGAACGCGGTAGAACGTGCTAATTGGATTTTGGTG AGAACGCGGTAGAACGTACTTATCACCTACAGTTTTATTTTGTTTTTCTTTTTGGTTTAATCTATCC AGCTTAGTACCGAGTGGGGGAAAGTGACTGGTGTGCCTAAAACCTTTTCTTTGATACTTTGTAAAAA TACATACAGATACAATGGCGAACGGTAACTTCAAGTTGTCTCAATTGCTCAATGTGGACGAGATGTC TGCTGAGCAGAGGAGTCATTTCTTTGACTTGATGCTGACTAAACCTGATTGTGAGATCGGGCAAATG ATGCAAAGAGTTGTTGTTGATAAAGTCGATGACATGATTAGAGAAAGAAAGACTAAAGATCCAGTGA TTGTTCATGAAGTTCTTTCTCAGAAGGAACAGAACAAGTTGATGGAAATTTATCCTGAATTCAATAT CGTGTTTAAAGACGACAAAAACATGGTTCATGGGTTTGCGGCTGCTGAGCGAAAACTACAAGCTTTA TTGCTTTTAGATAGAGTTCCTGCTCTGCAAGAGGTGGATGACATCGGTGGTCAATGGTCGTTTTGGG TAACTAGAGGTGAGAAAAGGATTCATTCCTGTTGTCCAAATCTAGATATTCGGGATGATCAGAGAGA AATTTCTCGACAGATATTTCTTACTGCTATTGGTGATCAAGCTAGAAGTGGTAAGAGACAGATGTCG GAGAATGAGCTGTGGATGTATGACCAATTTCGTGAAAATATTGCTGCGCCTAACGCGGTTAGGTGCA ATAATACATATCAGGGTTGTACATGTAGGGGTTTTTCTGATGGTAAGAAGAAAGGCGCGCAGTATGC GATAGCTCTTCACAGCCTGTATGACTTCAAGTTGAAAGACTTGATGGCTACTATGGTTGAGAAGAAA ACTAAAGTGGTTCATGCTGCTATGCTTTTTGCTCCTGAAAGTATGTTAGTGGACGAAGGTCCATTAC CTTCTGTTGACGGTTACTACATGAAGAAGAACGGGAAGATCTATTTCGGTTTTGAGAAAGATCCTTC CTTTTCTTACATTCATGACTGGGAAGAGTACAAGAAGTATCTACTGGGGAAGCCAGTGAGTTACCAA GGGAATGTGTTCTACTTCGAACCGTGGCAGGTGAGAGGAGACACAATGCTTTTTTCGATCTACAGGA TAGCTGGAGTTCCGAGGAGGTCTCTATCATCGCAAGAGTACTACCGAAGAATATATATCAGTAGATG GGAAAGCATGGTTGTTGTCCCAATTTTCGATCTGGTCGAATCAACGCGAGAGTTGGTCAAGAAAGAC CTGTTTGTAGAGAAACAATTCATGGACAAGTGTTTGGATTACATAGCTAGGTTATCTGACCAGCAGC TGACCATAAGCAATGTTAAATCATACTTGAGTTCAAATAATTGGGTCTTATTCATAAACGGGGCGGC CGTGAAGAACAAGCAAAGTGTAGATTCTCGAGATTTACAGTTGTTGGCTCAAACTTTGCTAGTGAAG GAACAAGTGGCGAGACCTGTCATGAGGGAGTTGCGTGAAGCAATTCTGACTGAGACGAAACCTATCA CGTCATTGACTGATGTGCTGGGTTTAATATCAAGAAAACTGTGGAAGCAGTTTGCTAACAAGATCGC AGTCGGCGGATTCGTTGGCATGGTTGGTACTCTAATTGGATTCTATCCAAAGAAGGTACTAACCTGG GCGAAGGACACACCAAATGGTCCAGAACTATGTTACGAGAACTCGCACAAAACCAAGGTGATAGTAT TTCTGAGTGTTGTGTATGCCATTGGAGGAATCACGCTTATGCGTCGAGACATCCGAGATGGACTGGT GAAAAAACTATGTGATATGTTTGATATCAAACGGGGGGCCCATGTCTTAGACGTTGAGAATCCGTGC CGCTATTATGAAATCAACGATTTCTTTAGCAGTCTGTATTCGGCATCTGAGTCCGGTGAGACCGTTT TACCAGATTTATCCGAGGTAAAAGCCAAGTCTGATAAGCTATTGCAGCAGAAGAAAGAAATCGCTGA CGAGTTTCTAAGTGCAAAATTCTCTAACTATTCTGGCAGTTCGGTGAGAACTTCTCCACCATCGGTG GTCGGTTCATCTCGAAGCGGACTGGGTCTGTTGTTGGAAGACAGTAACGTGCTGACCCAAGCTAGAG TTGGAGTTTCAAGAAAGGTAGACGATGAGGAGATCATGGAGCAGTTTCTGAGTGGTCTTATTGACAC TGAAGCAGAAATTGACGAGGTTGTTTCAGCCTTTTCAGCTGAATGTGAAAGAGGGGAAACAAGCGGT ACAAAGGTGTTGTGTAAACCTTTAACGCCACCAGGATTTGAGAACGTGTTGCCAGCTGTCAAACCTT TGGTCAGCAAAGGAAAAACGGTCAAACGTGTCGATTACTTCCAAGTGATGGGAGGTGAGAGATTACC AAAAAGGCCGGTTGTCAGTGGAGACGATTCTGTGGACGCTAGAAGAGAGTTTCTGTACTACTTAGAT GCGGAGAGAGTCGCTCAAAATGATGAAATTATGTCTCTGTATCGTGACTATTCGAGAGGAGTTATTC GAACTGGAGGTCAGAATTACCCGCACGGACTGGGAGTGTGGGATGTGGAGATGAAGAACTGGTGCAT ACGTCCAGTGGTCACTGAACATGCTTATGTGTTCCAACCAGACAAACGTATGGATGATTGGTCGGGA TACTTAGAAGTGGCTGTTTGGGAACGAGGTATGTTGGTCAACGACTTCGCGGTCGAAAGGATGAGTG ATTATGTCATAGTTTGCGATCAGACGTATCTTTGCAATAACAGGTAATAATCCTCTCTCTTGATATT TTTAAATTATAGAATTAATTAGTTTACTTTATTCTTTACTATATGATTTAAATAGTTTAATCTTGTT TTTGAGTAAACTATTCGATTTTGATATTTGTATTCGTCCTACAAAGTTGGAAATACTGATGATATTT TCTTTTGAACGTGATACCTACCAATACTAATCTTACGGAATCTTTTAATAGAGCACTAATCAACATG GAACTAAAGACCAATTCTTAAGTGTCTCTGTTGTACAGTTCATTTTAGTAGTGCGTTTAAGTATTAT TATCTCCCTTCATGCGGGGCAATTATGTAGATTAAAATCGAAATTATATAAAATTTACATAAGTCTA AGTCTAGGGTCTCCAGCTAATTGTTATTTTTTTAACGATGTTGACTAAAGCAATAACGACGTTGACT TGTGTTAAACAGGTTGATCTTGGACAATTTAAGTGCCCTGGATCTAGGACCAGTTAACTGTTCTTTT GAATTAGTTGACGGTGTACCTGGTTGTGGTAAGTCGACAATGATTGTCAACTCAGCTAATCCTTGTG TCGATGTGGTTCTCTCTACTGGGAGAGCAGCAACCGACGACTTGATCGAGAGATTCGCGAGCAAAGG TTTTCCATGCAAATTGAAAAGGAGAGTGAAGACGGTTGATTCTTTTTTGATGCATTGTGTCGATGGT TCTTTAACCGGAGACGTGTTGCATTTCGACGAAGCTCTCATGGCCCATGCTGGTATGGTGTACTTTT GCGCTCAGATAGCTGGTGCTAAACGATGTATCTGTCAAGGAGATCAGAATCAAATTTCTTTCAAGCC TAGGGTATCTCAAGTTGATTTGAGGTTTTCTAGTCTGGTCGGAAAGTTTGACATTGTTACAGAAAAA AGAGAAACTTACAGAAGTCCAGCAGATGTGGCTGCCGTATTGAACAAGTACTATACTGGAGATGTCA GAACACATAACGCGACTGCTAATTCGATGACGGTGAGGAAGATTGTGTCTAAAGAACAGGTTTCTTT GAAGCCTGGTGCTCAGTACATAACTTTCCTTCAGTCTGAGAAGAAGGAGTTGGTAAATTTGTTGGCA TTGAGGAAAGTGGCAGCTAAAGTGAGTACAGTACACGAGTCGCAAGGAGAGACATTCAAAGATGTAG TCCTAGTCAGGACGAAACCTACGGATGACTCAATCGCTAGAGGTCGGGAGTACTTAATCGTGGCATT GTCGCGTCACACACAATCACTTGTGTATGAAACTGTGAAAGAGGACGATGTAAGCAAAGAGATCAGG GAAAGTGCCGCGCTTACGAAGGCGGCTTTGGCAAGATTTTTTGTTACTGAGACCGTCTTATGACGGT TTCGGTCTAGGTTTGATGTCTTTAGACATCATGAAGGGCCTTGCGCCGTTCCAGATTCAGGTACGAT TACGGACTTGGAGATGTGGTACGACGCTTTGTTTCCGGGAAATTCGTTAAGAGACTCAAGCCTAGAC GGGTATTTGGTGGCAACGACTGATTGCAATTTGCGATTAGACAATGTTACGATCAAAAGTGGAAACT GGAAAGACAAGTTTGCTGAAAAAGAAACGTTTCTGAAACCGGTTATTCGTACTGCTATGCCTGACAA AAGGAAGACTACTCAGTTGGAGAGTTTGTTAGCATTGCAGAAAAGGAACCAAGCGGCACCCGATCTA CAAGAAAATGTGCACGCGACAGTTCTAATCGAAGAGACGATGAAGAAGCTGAAATCTGTTGTCTACG ATGTGGGAAAAATTCGGGCTGATCCTATTGTCAATAGAGCTCAAATGGAGAGATGGTGGAGAAATCA AAGCACAGCGGTACAGGCTAAGGTAGTAGCAGATGTGAGAGAGTTACATGAAATAGACTATTCGTCT TACATGTATATGATCAAATCTGACGTGAAACCTAAGACTGATTTAACACCGCAATTTGAATACTCAG CTCTACAGACTGTTGTGTATCACGAGAAGTTGATCAACTCGTTGTTCGGTCCAATTTTCAAAGAAAT TAATGAACGCAAGTTGGATGCTATGCAACCACATTTTGTGTTCAACACGAGAATGACATCGAGTGAT TTAAACGATCGAGTGAAGTTCTTAAATACGGAAGCGGCTTACGACTTTGTTGAGATAGACATGTCTA AATTCGACAAGTCGGCAAATCGCTTCCATTTACAACTGCAGCTGGAGATTTACAGGTTATTTGGGCT GGATGAGTGGGCGGCCTTCCTTTGGGAGGTGTCGCACACTCAAACTACTGTGAGAGATATTCAAAAT GGTATGATGGCGCATATTTGGTACCAACAAAAGAGTGGAGATGCTGATACTTATAATGCAAATTCAG ATAGAACACTGTGTGCACTCTTGTCTGAATTACCATTGGAGAAAGCAGTCATGGTTACATATGGAGG AGATGACTCACTGATTGCGTTTCCTAGAGGAACGCAGTTTGTTGATCCGTGTCCAAAGTTGGCTACT AAGTGGAATTTCGAGTGCAAGATTTTTAAGTACGATGTCCCAATGTTTTGTGGGAAGTTCTTGCTTA AGACGTCATCGTGTTACGAGTTCGTGCCAGATCCGGTAAAAGTTCTGACGAAGTTGGGGAAAAAGAG TATAAAGGATGTGCAACATTTAGCCGAGATCTACATCTCGCTGAATGATTCCAATAGAGCTCTTGGG AACTACATGGTGGTATCCAAACTGTCCGAGTCTGTTTCAGACCGGTATTTGTACAAAGGTGATTCTG TTCATGCGCTTTGTGCGCTATGGAAGCATATTAAGAGTTTTACAGCTCTGTGTACATTATTCCGAGA CGAAAACGATAAGGAATTGAACCCGGCTAAGGTTGATTGGAAGAAGGCACAGAGAGCTGTGTCAAAC TTTTACGACTGGTAATATGGAAGACAAGTCATTGGTCACCTTGAAGAAGAAGACT GGCGCGCCACGT GTTAATTAA CTGATTCGACTAGGCGCCTCAATGTGGAAGAACTGAACAGTTCGGATTACATTGAAGG CGATTTTACCGATCAAGAGGTTTTCGGTGAGTTCATGTCTTTGAAACAAGTGGAGATGAAGACGATT GAGGCGAAGTACGATGGTCCTTACAGACCAGCTACTACTAGACCTAAGTCATTATTGTCAAGTGAAG ATGTTAAGAGAGCGTCTAATAAGAAAAACTCGTCTTAATGCATAAAGAAATTTATTGTCAATATGAC GTGTGTACTCAAGGGTTGTGTGAATGAAGTCACTGTTCTTGGTCACGAGACGTGTAGTATCGGTCAT GCTAACAAATTGCGAAAGCAAGTTGCTGACATGGTTGGTGTCACACGTAGGTGTGCGGAAAATAATT GTGGATGGTTTGTCTGTGTTGTTATCAATGATTTTACTTTTGATGTGTATAATTGTTGTGGCCGTAG TCACCTTGAAAAGTGTCGTAAACGTGTTGAAACAAGAAATCGAGAAATTTGGAAACAAATTCGACGA AATCAAGCTGAAAACATGTCTGCGACAGCTAAAAAGTCTCATAATTCGAAGACCTCTAAGAAGAAAT TCAAAGAGGACAGAGAATTTGGGACACCAAAAAGATTTTTAAGAGATGATGTTCCTTTCGGGATTGA TCGTTTGTTTGCTTTTTGATTTTATTTTATATTGTTATCTGTTTCTGTGTATAGACTGTTTGAGATT GGCGCTTGGCCGACTCATTGTCTTACCATAGGGGAACGGACTTTGTTTGTGTTGTTATTTTATTTGT ATTTTATTAAAATTCTCAATGATCTGAAAAGGCCTCGAGGCTAAGAGATTATTGGGGGGTGAGTAAG TACTTTTAAAGTGATGATGGTTACAAAGGCAAAAGGGGTAAAACCCCTCGCCTACGTAAGCGTTATT ACGCCCGgatcccccggggagctcgaattcgctgaaatcaccagtctctctctacaaatctatctct ctctattttttccataaataatgtgtgagtagtttcccgataagggaaattagggttcttatagggt ttcgctcatgtgttgagcatataagaaacccttagtatgtatttgtatttgtaaaatacttctatta tcaataaaatttctaattcctaaaaccaaaatccagtactaaaatccagatctcctaaagtccctat agatctttgtcgtgaatataaaccagacacgagacgactaaacctggagcccagacgccgttcgaag ctagaagtaccgcttaggcaggaggccgttagggaaaagatgctaaggcagggttggttacgttgac tcccccgtaggtttggtttaaatatgatgaagtggacggaaggaaggaggaagacaaggaaggataa ggttgcaggccctgtgcaaggtaagaagatggaaatttgatagaggtacgctactatacttatacta tacgctaagggaatgcttgtatttataccctatacaccotaataaccccttatcaatttaagaaata atccgcataagcccccgcttaaaaattggtatcagagccatgaataggtctatgaccaaaactcaag aggataaaacctcaccaaaatacgaaagagttcttaactctaaagataaaagatctttcaagatcaa aactagttccctcacaccggagcatgcgatatcctcgacctgcaggcatgcaagcttggcgtaatca tggtcatagctgtttcctgtgtgaaattgttatccgctcacaattccacacaacatacgagccggaa gcataaagtgtaaagcctggggtgcctaatgagtgagctaactcacattaattgcgttgcgctcact gcccgctttccagtcgggaaacctgtcgtgccagctgcattaatgaatcggccaacgcgcggggaga ggcggtttgcgtattgggccaaagacaaaagggcgacattcaaccgattgagggagggaaggtaaat attgacggaaattattcattaaaggtgaattatcaccgtcaccgacttgagccatttgggaattaga gccagcaaaatcaccagtagcaccattaccattagcaaggccggaaacgtcaccaatgaaaccatcg atagcagcaccgtaatcagtagcgacagaatcaagtttgcctttagcgtcagactgtagcgcgtttt catcggcattttcggtcatagcccccttattagcgtttgccatcttttcataatcaaaatcaccgga accagagccaccaccggaaccgcctccctcagagccgccaccctcagaaccgccaccctcagagcca ccaccctcagagccgccaccagaaccaccaccagagccgccgccagcattgacaggaggcccgatct agtaacatagatgacaccgcgcgcgataatttatcctagtttgcgcgctatattttgttttctatcg cgtattaaatgtataattgcgggactctaatcataaaaacccatctcataaataacgtcatgcatta catgttaattattacatgcttaacgtaattcaacagaaattatatgataatcatcgcaagaccggca acaggattcaatcttaagaaactttattgccaaatgtttgaacgatcggggatcatccgggtctgtg gcgggaactccacgaaaatatccgaacgcagcaagatatcgcggtgcatctcggtcttgcctgggca gtcgccgccgacgccgttgatgtggacgccgggcccgatcatattgtcgctcaggatcgtggcgttg tgcttgtcggccgttgctgtcgtaatgatatcggcaccttcgaccgcctgttccgcagagatcccgt gggcgaagaactccagcatgagatccccgcgctggaggatcatccagccggcgtcccggaaaacgat tccgaagcccaacctttcatagaaggcggcggtggaatcgaaatctcgtgatggcaggttgggcgtc gcttggtcggtcatttcgaaccccagagtcccgctcagaagaactcgtcaagaaggcgatagaaggc gatgcgctgcgaatcgggagcggcgataccgtaaagcacgaggaagcggtcagcccattcgccgcca agctcttcagcaatatcacgggtagccaacgctatgtcctgatagcggtccgccacacccagccggc cacagtcgatgaatccagaaaagcggccattttccaccatgatattcggcaagcaggcatcgccatg ggtcacgacgagatcatcgccgtcgggcatgcgcgccttgagcctggcgaacagttcggctggcgcg agcccctgatgctcttcgtccagatcatcctgatcgacaagaccggcttccatccgagtacgtgctc gctcgatgcgatgtttcgcttggtggtcgaatgggcaggtagccggatcaagcgtatgcagccgccg cattgcatcagccatgatggatactttctcggcaggagcaaggtgagatgacaggagatcctgcccc ggcacttcgcccaatagcagccagtcccttcccgcttcagtgacaacgtcgagcacagctgcgcaag gaacgcccgtcgtggccagccacgatagccgcgctgcctcgtcctgcagttcattcagggcaccgga caggtcggtcttgacaaaaagaaccgggcgcccctgcgctgacagccggaacacggcggcatcagag cagccgattgtctgttgtgcccagtcatagccgaatagcctctccacccaagcggccggagaacctg cgtgcaatccatcttgttcaatcatgcgaaacgatccagatccggtgcagattatttggattgagag tgaatatgagactctaattggataccgaggggaatttatggaacgtcagtggagcatttttgacaag aaatatttgctagctgatagtgaccttaggcgacttttgaacgcgcaataatggtttctgacgtatg tgcttagctcattaaactccagaaacccgcggctgagtggctccttcaacgttgcggttctgtcagt tccaaacgtaaaacggcttgtcccgcgtcatcggcgggggtcataacgtgactcccttaattctccg ctcatgatcagattgtcgtttcccgccttcagtttaaactatcagtgtttgacaggatatattggcg ggtaaacctaagagaaaagagcgtttattagaataatcggatatttaaaagggcgtgaaaaggttta tccgttcgtccatttgtatgtgcatgccaaccacagggttccccagatctggcgccggccagcgaga cgagcaagattggccgccgcccgaaacgatccgacagcgcgcccagcacaggtgcgcaggcaaattg caccaacgcatacagcgccagcagaatgccatagtgggcggtgacgtcgttcgagtgaaccagatcg cgcaggaggcccggcagcaccggcataatcaggccgatgccgacagcgtcgagcgcgacagtgctca gaattacgatcaggggtatgttgggtttcacgtctggcctccggaccagcctccgctggtccgattg aacgcgcggattctttatcactgataagttggtggacatattatgtttatcagtgataaagtgtcaa gcatgacaaagttgcagccgaatacagtgatccgtgccgccctggacctgttgaacgaggtcggcgt agacggtctgacgacacgcaaactggcggaacggttgggggttcagcagccggcgctttactggcac ttcaggaacaagcgggcgctgctcgacgcactggccgaagccatgctggcggagaatcatacgcatt cggtgccgagagccgacgacgactggcgctcatttctgatcgggaatgcccgcagcttcaggcaggc gctgctcgcctaccgcgatggcgcgcgcatccatgccggcacgcgaccgggcgcaccgcagatggaa acggccgacgcgcagcttcgcttcctctgcgaggcgggtttttcggccggggacgccgtcaatgcgc tgatgacaatcagctacttcactgttggggccgtgcttgaggagcaggccggcgacagcgatgccgg cgagcgcggcggcaccgttgaacaggctccgctctcgccgctgttgcgggccgcgatagacgccttc gacgaagccggtccggacgcagcgttcgagcagggactcgcggtgattgtcgatggattggcgaaaa ggaggctcgttgtcaggaacgttgaaggaccgagaaagggtgacgattgatcaggaccgctgccgga gcgcaacccactcactacagcagagccatgtagacaacatcccctccccctttccaccgcgtcagac gcccgtagcagcccgctacgggctttttcatgccctgccctagcgtccaagcctcacggccgcgctc ggcctctctggcggccttctggcgctcttccgcttcctcgctcactgactcgctgcgctcggtcgtt cggctgcggcgagcggtatcagctcactcaaaggcggtaatacggttatccacagaatcaggggata acgcaggaaagaacatgtgagcaaaaggccagcaaaaggccaggaaccgtaaaaaggccgcgttgct ggcgtttttccataggctccgcccccctgacgagcatcacaaaaatcgacgctcaagtcagaggtgg cgaaacccgacaggactataaagataccaggcgtttccccctggaagctccctcgtgcgctctcctg ttccgaccctgccgcttaccggatacctgtccgcctttctcccttcgggaagcgtggcgcttttccg ctgcataaccctgcttcggggtcattatagcgattttttcggtatatccatcctttttcgcacgata tacaggattttgccaaagggttcgtgtagactttccttggtgtatccaacggcgtcagccgggcagg ataggtgaagtaggcccacccgcgagcgggtgttccttcttcactgtcccttattcgcacctggcgg tgctcaacgggaatcctgctctgcgaggctggccggctaccgccggcgtaacagatgagggcaagcg gatggctgatgaaaccaagccaaccaggaagggcagcccacctatcaaggtgtactgccttccagac gaacgaagagcgattgaggaaaaggcggcggcggccggcatgagcctgtcggcctacctgctggccg tcggccagggctacaaaatcacgggcgtcgtggactatgagcacgtccgcgagctggcccgcatcaa tggcgacctgggccgcctgggcggcctgctgaaactctggctcaccgacgacccgcgcacggcgcgg ttcggtgatgccacgatcctcgccctgctggcgaagatcgaagagaagcaggacgagcttggcaagg tcatgatgggcgtggtccgcccgagggcagagccatgacttttttagccgctaaaacggccgggggg tgcgcgtgattgccaagcacgtccccatgcgctccatcaagaagagcgacttcgcggagctggtgaa gtacatcaccgacgagcaaggcaagaccgagcgcctttgcgacgctcaccgggctggttgccctcgc cgctgggctggcggccgtctatggccctgcaaacgcgccagaaacgccgtcgaagccgtgtgcgaga caccgcggccgccggcgttgtggatacctcgcggaaaacttggccctcactgacagatgaggggcgg acgttgacacttgaggggccgactcacccggcgcggcgttgacagatgaggggcaggctcgatttcg gccggcgacgtggagctggccagcctcgcaaatcggcgaaaacgcctgattttacgcgagtttccca cagatgatgtggacaagcctggggataagtgccctgcggtattgacacttgaggggcgcgactactg acagatgaggggcgcgatccttgacacttgaggggcagagtgctgacagatgaggggcgcacctatt gacatttgaggggctgtccacaggcagaaaatccagcatttgcaagggtttccgcccgtttttcggc caccgctaacctgtcttttaacctgcttttaaaccaatatttataaaccttgtttttaaccagggct gcgccctgtgcgcgtgaccgcgcacgccgaaggggggtgcccccccttctcgaaccctcccggcccg ctaacgcgggcctcccatccccccaggggctgcgcccctcggccgcgaacggcctcaccccaaaaat ggcagcgctggcagtccttgccattgccgggatcggggcagtaacgggatgggcgatcagcccgagc gcgacgcccggaagcattgacgtgccgcaggtgctggcatcgacattcagcgaccaggtgccgggca gtgagggcggcggcctgggtggcggcctgcccttcacttcggccgtcggggcattcacggacttcat ggcggggccggcaatttttaccttgggcattcttggcatagtggtcgcgggtgccgtgctcgtgttc gggggtgcgataaacccagcgaaccatttgaggtgataggtaagattataccgaggtatgaaaacga gaattggacctttacagaattactctatgaagcgccatatttaaaaagctaccaagacgaagaggat gaagaggatgaggaggcagattgccttgaatatattgacaatactgataagataatatatcttttat atagaagatatcgccgtatgtaaggatttcagggggcaaggcataggcagcgcgcttatcaatatat ctatagaatgggcaaagcataaaaacttgcatggactaatgcttgaaacccaggacaataaccttat agcttgtaaattctatcataattgggtaatgactccaacttattgatagtgttttatgttcagataa tgcccgatgactttgtcatgcagctccaccgattttgagaacgacagcgacttccgtcccagccgtg ccaggtgctgcctcagattcaggttatgccgctcaattcgctgcgtatatcgcttgctgattacgtg cagctttcccttcaggcgggattcatacagcggccagccatccgtcatccatatcaccacgtcaaag ggtgacagcaggctcataagacgccccagcgtcgccatagtgcgttcaccgaatacgtgcgcaacaa ccgtcttccggagactgtcatacgcgtaaaacagccagcgctggcgcgatttagccccgacatagcc ccactgttcgtccatttccgcgcagacgatgacgtcactgcccggctgtatgcgcgaggttaccgac tgcggcctgagttttttaagtgacgtaaaatcgtgttgaggccaacgcccataatgcgggctgttgc ccggcatccaacgccattcatggccatatcaatgattttctggtgcgtaccgggttgagaagcggtg taagtgaactgcagttgccatgttttacggcagtgagagcagagatagcgctgatgtccggcggtgc ttttgccgttacgcaccaccccgtcagtagctgaacaggagggacagctgatagacacagaagccac tggagcacctcaaaaacaccatcatacactaaatcagtaagttggcagcatcacccataattgtggt ttcaaaatcggctccgtcgatactatgttatacgccaactttgaaaacaactttgaaaaagctgttt tctggtatttaaggttttagaatgcaaggaacagtgaattggagttcgtcttgttataattagcttc ttggggtatctttaaatactgtagaaaagaggaaggaaataataaatggctaaaatgagaatatcac cggaattgaaaaaactgatcgaaaaataccgctgcgtaaaagatacggaaggaatgtctcctgctaa ggtatataagctggtgggagaaaatgaaaacctatatttaaaaatgacggacagccggtataaaggg accacctatgatgtggaacgggaaaaggacatgatgctatggctggaaggaaagctgcctgttccaa aggtcctgcactttgaacggcatgatggctggagcaatctgctcatgagtgaggccgatggcgtcct ttgctcggaagagtatgaagatgaacaaagccctgaaaagattatcgagctgtatgcggagtgcatc aggctctttcactccatcgacatatcggattgtccctatacgaatagcttagacagccgcttagccg aattggattacttactgaataacgatctggccgatgtggattgcgaaaactgggaagaagacactcc atttaaagatccgcgcgagctgtatgattttttaaagacggaaaagcccgaagaggaacttgtcttt tcccacggcgacctgggagacagcaacatctttgtgaaagatggcaaagtaagtggctttattgatc ttgggagaagcggcagggcggacaagtggtatgacattgccttctgcgtccggtcgatcagggagga tatcggggaagaacagtatgtcgagctattttttgacttactggggatcaagcctgattgggagaaa ataaaatattatattttactggatgaattgttttagtacctagatgtggcgcaacgatgccggcgac aagcaggagcgcaccgacttcttccgcatcaagtgttttggctctcaggccgaggcccacggcaagt atttgggcaaggggtcgctggtattcgtgcagggcaagattcggaataccaagtacgagaaggacgg ccagacggtctacgggaccgacttcattgccgataaggtggattatctggacaccaaggcaccaggc gggtcaaatcaggaataagggcacattgccccggcgtgagtcggggcaatcccgcaaggagggtgaa tgaatcggacgtttgaccggaaggcatacaggcaagaactgatcgacgcggggttttccgccgagga tgccgaaaccatcgcaagccgcaccgtcatgcgtgcgccccgcgaaaccttccagtccgtcggctcg atggtccagcaagctacggccaagatcgagcgcgacagcgtgcaactggctccccctgccctgcccg cgccatcggccgccgtggagcgttcgcgtcgtctcgaacaggaggcggcaggtttggcgaagtcgat gaccatcgacacgcgaggaactatgacgaccaagaagcgaaaaaccgccggcgaggacctggcaaaa caggtcagcgaggccaagcaggccgcgttgctgaaacacacgaagcagcagatcaaggaaatgcagc tttccttgttcgatattgcgccgtggccggacacgatgcgagcgatgccaaacgacacggcccgctc tgccctgttcaccacgcgcaacaagaaaatcccgcgcgaggcgctgcaaaacaaggtcattttccac gtcaacaaggacgtgaagatcacctacaccggcgtcgagctgcgggccgacgatgacgaactggtgt ggcagcaggtgttggagtacgcgaagcgcacccctatcggcgagccgatcaccttcacgttctacga gctttgccaggacctgggctggtcgatcaatggccggtattacacgaaggccgaggaatgcctgtcg cgcctacaggcgacggcgatgggcttcacgtccgaccgcgttgggcacctggaatcggtgtcgctgc tgcaccgcttccgcgtcctggaccgtggcaagaaaacgtcccgttgccaggtcctgatcgacgagga aatcgtcgtgctgtttgctggcgaccactacacgaaattcatatgggagaagtaccgcaagctgtcg ccgacggcccgacggatgttcgactatttcagctcgcaccgggagccgtacccgctcaagctggaaa ccttccgcctcatgtgcggatcggattccacccgcgtgaagaagtggcgcgagcaggtcggcgaagc ctgcgaagagttgcgaggcagcggcctggtggaacacgcctgggtcaatgatgacctggtgcattgc aaacgctagggccttgtggggtcagttccggctgggggttcagcagccagcgctttactggcatttc aggaacaagcgggcactgctcgacgcacttgcttcgctcagtatcgctcgggacgcacggcgcgctc tacgaactgccgataaacagaggattaaaattgacaattgtgattaaggctcagattcgacggcttg gagcggccgacgtgcaggatttccgcgagatccgattgtcggccctgaagaaagctccagagatgtt cgggtccgtttacgagcacgaggagaaaaagcccatggaggcgttcgctgaacggttgcgagatgcc gtggcattcggcgcctacatcgacggcgagatcattgggctgtcggtcttcaaacaggaggacggcc ccaaggacgctcacaaggcgcatctgtccggcgttttcgtggagcccgaacagcgaggccgaggggt cgccggtatgctgctgcgggcgttgccggcgggtttattgctcgtgatgatcgtccgacagattcca acgggaatctggtggatgcgcatcttcatcctcggcgcacttaatatttcgctattctggagcttgt tgtttatttcggtctaccgcctgccgggcggggtcgcggcgacggtaggcgctgtgcagccgctgat ggtcgtgttcatctctgccgctctgctaggtagcccgatacgattgatggcggtcctgggggctatt tgcggaactgcgggcgtggcgctgttggtgttgacaccaaacgcagcgctagatcctgtcggcgtcg cagcgggcctggcgggggcggtttccatggcgttcggaaccgtgctgacccgcaagtggcaacctcc cgtgcctctgctcacctttaccgcctggcaactggcggccggaggacttctgctcgttccagtagct ttagtgtttgatccgccaatcccgatgcctacaggaaccaatgttctcggcctggcgtggctcggcc tgatcggagcgggtttaacctacttcctttggttccgggggatctcgcgactcgaacctacagttgt ttccttactgggctttctcagccccagatctggggtcgatcagccggggatgcatcaggccgacagt cggaacttcgggtccccgacctgtaccattcggtgagcaatggataggggagttgatatcgtcaacg ttcacttctaaagaaatagcgccactcagcttcctcagcggctttatccagcgatttcctattatgt cggcatagttctcaagatcgacagcctgtcacggttaagcgagaaatgaataagaaggctgataatt cggatctctgcgagggagatgatatttgatcacaggcagcaacgctctgtcatcgttacaatcaaca tgctaccctccgcgagatcatccgtgtttcaaacccggcagcttagttgccgttcttccgaatagca tcggtaacatgagcaaagtctgccgccttacaacggctctcccgctgacgccgtcccggactgatgg gctgcctgtatcgagtggtgattttgtgccgagctgccggtcggggagctgttggctggctggtggc aggatatattgtggtgtaaacaaattgacgcttagacaacttaataacacattgcggacgtttttaa tgtactggggtggtttttcttttcaccagtgagacgggcaacagctgattgcccttcaccgcctggc cctgagagagttgcagcaagcggtccacgctggtttgccccagcaggcgaaaatcctgtttgatggt ggttccgaaatcggcaaaatcccttataaatcaaaagaatagcccgagatagggttgagtgttgttc cagtttggaacaagagtccactattaaagaacgtggactccaacgtcaaagggcgaaaaaccgtcta tcagggcgatggcccactacgtgaaccatcacccaaatcaagttttttggggtcgaggtgccgtaaa gcactaaatcggaaccctaaagggagcccccgatttagagcttgacggggaaagccggcgaacgtgg cgagaaaggaagggaagaaagcgaaaggagcgggcgccattcaggctgcgcaactgttgggaagggc gatcggtgcgggcctcttcgctattacgccagctggcgaaagggggatgtgctgcaaggcgattaag ttgggtaacgccagggttttcccagtcacgacgttgtaaaacgacggccagtgaattcgagctcggt acccccc SEQ ID NO 3 - pBTAΔMPΔ16K GATCCATAAAACATTTCAATCCTTTGAACGCGGTAGAACGTGCTAATTGGATTTTGGTGAGAACGCG GTAGAACGTACTTATCACCTACAGTTTTATTTTGTTTTTCTTTTTGGTTTAATCTATCCAGCTTAGT ACCGAGTGGGGGAAAGTGACTGGTGTGCCTAAAACCTTTTCTTTGATACTTTGTAAAAATACATACA GATACAATGGCGAACGGTAACTTCAAGTTGTCTCAATTGCTCAATGTGGACGAGATGTCTGCTGAGC AGAGGAGTCATTTCTTTGACTTGATGCTGACTAAACCTGATTGTGAGATCGGGCAAATGATGCAAAG AGTTGTTGTTGATAAAGTCGATGACATGATTAGAGAAAGAAAGACTAAAGATCCAGTGATTGTTCAT GAAGTTCTTTCTCAGAAGGAACAGAACAAGTTGATGGAAATTTATCCTGAATTCAATATCGTGTTTA AAGACGACAAAAACATGGTTCATGGGTTTGCGGCTGCTGAGCGAAAACTACAAGCTTTATTGCTTTT AGATAGAGTTCCTGCTCTGCAAGAGGTGGATGACATCGGTGGTCAATGGTCGTTTTGGGTAACTAGA GGTGAGAAAAGGATTCATTCCTGTTGTCCAAATCTAGATATTCGGGATGATCAGAGAGAAATTTCTC GACAGATATTTCTTACTGCTATTGGTGATCAAGCTAGAAGTGGTAAGAGACAGATGTCGGAGAATGA GCTGTGGATGTATGACCAATTTCGTGAAAATATTGCTGCGCCTAACGCGGTTAGGTGCAATAATACA TATCAGGGTTGTACATGTAGGGGTTTTTCTGATGGTAAGAAGAAAGGCGCGCAGTATGCGATAGCTC TTCACAGCCTGTATGACTTCAAGTTGAAAGACTTGATGGCTACTATGGTTGAGAAGAAAACTAAAGT GGTTCATGCTGCTATGCTTTTTGCTCCTGAAAGTATGTTAGTGGACGAAGGTCCATTACCTTCTGTT GACGGTTACTACATGAAGAAGAACGGGAAGATCTATTTCGGTTTTGAGAAAGATCCTTCCTTTTCTT ACATTCATGACTGGGAAGAGTACAAGAAGTATCTACTGGGGAAGCCAGTGAGTTACCAAGGGAATGT GTTCTACTTCGAACCGTGGCAGGTGAGAGGAGACACAATGCTTTTTTCGATCTACAGGATAGCTGGA GTTCCGAGGAGGTCTCTATCATCGCAAGAGTACTACCGAAGAATATATATCAGTAGATGGGAAAGCA TGGTTGTTGTCCCAATTTTCGATCTGGTCGAATCAACGCGAGAGTTGGTCAAGAAAGACCTGTTTGT AGAGAAACAATTCATGGACAAGTGTTTGGATTACATAGCTAGGTTATCTGACCAGCAGCTGACCATA AGCAATGTTAAATCATACTTGAGTTCAAATAATTGGGTCTTATTCATAAACGGGGCGGCCGTGAAGA ACAAGCAAAGTGTAGATTCTCGAGATTTACAGTTGTTGGCTCAAACTTTGCTAGTGAAGGAACAAGT GGCGAGACCTGTCATGAGGGAGTTGCGTGAAGCAATTCTGACTGAGACGAAACCTATCACGTCATTG ACTGATGTGCTGGGTTTAATATCAAGAAAACTGTGGAAGCAGTTTGCTAACAAGATCGCAGTCGGCG GATTCGTTGGCATGGTTGGTACTCTAATTGGATTCTATCCAAAGAAGGTACTAACCTGGGCGAAGGA CACACCAAATGGTCCAGAACTATGTTACGAGAACTCGCACAAAACCAAGGTGATAGTATTTCTGAGT GTTGTGTATGCCATTGGAGGAATCACGCTTATGCGTCGAGACATCCGAGATGGACTGGTGAAAAAAC TATGTGATATGTTTGATATCAAACGGGGGGCCCATGTCTTAGACGTTGAGAATCCGTGCCGCTATTA TGAAATCAACGATTTCTTTAGCAGTCTGTATTCGGCATCTGAGTCCGGTGAGACCGTTTTACCAGAT TTATCCGAGGTAAAAGCCAAGTCTGATAAGCTATTGCAGCAGAAGAAAGAAATCGCTGACGAGTTTC TAAGTGCAAAATTCTCTAACTATTCTGGCAGTTCGGTGAGAACTTCTCCACCATCGGTGGTCGGTTC ATCTCGAAGCGGACTGGGTCTGTTGTTGGAAGACAGTAACGTGCTGACCCAAGCTAGAGTTGGAGTT TCAAGAAAGGTAGACGATGAGGAGATCATGGAGCAGTTTCTGAGTGGTCTTATTGACACTGAAGCAG AAATTGACGAGGTTGTTTCAGCCTTTTCAGCTGAATGTGAAAGAGGGGAAACAAGCGGTACAAAGGT GTTGTGTAAACCTTTAACGCCACCAGGATTTGAGAACGTGTTGCCAGCTGTCAAACCTTTGGTCAGC AAAGGAAAAACGGTCAAACGTGTCGATTACTTCCAAGTGATGGGAGGTGAGAGATTACCAAAAAGGC CGGTTGTCAGTGGAGACGATTCTGTGGACGCTAGAAGAGAGTTTCTGTACTACTTAGATGCGGAGAG AGTCGCTCAAAATGATGAAATTATGTCTCTGTATCGTGACTATTCGAGAGGAGTTATTCGAACTGGA GGTCAGAATTACCCGCACGGACTGGGAGTGTGGGATGTGGAGATGAAGAACTGGTGCATACGTCCAG TGGTCACTGAACATGCTTATGTGTTCCAACCAGACAAACGTATGGATGATTGGTCGGGATACTTAGA AGTGGCTGTTTGGGAACGAGGTATGTTGGTCAACGACTTCGCGGTCGAAAGGATGAGTGATTATGTC ATAGTTTGCGATCAGACGTATCTTTGCAATAACAGGTAATAATCCTCTCTCTTGATATTTTTAAATT ATAGAATTAATTAGTTTACTTTATTCTTTACTATATGATTTAAATAGTTTAATCTTGTTTTTGAGTA AACTATTCGATTTTGATATTTGTATTCGTCCTACAAAGTTGGAAATACTGATGATATTTTCTTTTGA ACGTGATACCTACCAATACTAATCTTACGGAATCTTTTAATAGAGCACTAATCAACATGGAACTAAA GACCAATTCTTAAGTGTCTCTGTTGTACAGTTCATTTTAGTAGTGCGTTTAAGTATTATTATCTCCC TTCATGCGGGGCAATTATGTAGATTAAAATCGAAATTATATAAAATTTACATAAGTCTAAGTCTAGG GTCTCCAGCTAATTGTTATTTTTTTAACGATGTTGACTAAAGCAATAACGACGTTGACTTGTGTTAA ACAGGTTGATCTTGGACAATTTAAGTGCCCTGGATCTAGGACCAGTTAACTGTTCTTTTGAATTAGT TGACGGTGTACCTGGTTGTGGTAAGTCGACAATGATTGTCAACTCAGCTAATCCTTGTGTCGATGTG GTTCTCTCTACTGGGAGAGCAGCAACCGACGACTTGATCGAGAGATTCGCGAGCAAAGGTTTTCCAT GCAAATTGAAAAGGAGAGTGAAGACGGTTGATTCTTTTTTGATGCATTGTGTCGATGGTTCTTTAAC CGGAGACGTGTTGCATTTCGACGAAGCTCTCATGGCCCATGCTGGTATGGTGTACTTTTGCGCTCAG ATAGCTGGTGCTAAACGATGTATCTGTCAAGGAGATCAGAATCAAATTTCTTTCAAGCCTAGGGTAT CTCAAGTTGATTTGAGGTTTTCTAGTCTGGTCGGAAAGTTTGACATTGTTACAGAAAAAAGAGAAAC TTACAGAAGTCCAGCAGATGTGGCTGCCGTATTGAACAAGTACTATACTGGAGATGTCAGAACACAT AACGCGACTGCTAATTCGATGACGGTGAGGAAGATTGTGTCTAAAGAACAGGTTTCTTTGAAGCCTG GTGCTCAGTACATAACTTTCCTTCAGTCTGAGAAGAAGGAGTTGGTAAATTTGTTGGCATTGAGGAA AGTGGCAGCTAAAGTGAGTACAGTACACGAGTCGCAAGGAGAGACATTCAAAGATGTAGTCCTAGTC AGGACGAAACCTACGGATGACTCAATCGCTAGAGGTCGGGAGTACTTAATCGTGGCATTGTCGCGTC ACACACAATCACTTGTGTATGAAACTGTGAAAGAGGACGATGTAAGCAAAGAGATCAGGGAAAGTGC CGCGCTTACGAAGGCGGCTTTGGCAAGATTTTTTGTTACTGAGACCGTCTTATGACGGTTTCGGTCT AGGTTTGATGTCTTTAGACATCATGAAGGGCCTTGCGCCGTTCCAGATTCAGGTACGATTACGGACT TGGAGATGTGGTACGACGCTTTGTTTCCGGGAAATTCGTTAAGAGACTCAAGCCTAGACGGGTATTT GGTGGCAACGACTGATTGCAATTTGCGATTAGACAATGTTACGATCAAAAGTGGAAACTGGAAAGAC AAGTTTGCTGAAAAAGAAACGTTTCTGAAACCGGTTATTCGTACTGCTATGCCTGACAAAAGGAAGA CTACTCAGTTGGAGAGTTTGTTAGCATTGCAGAAAAGGAACCAAGCGGCACCCGATCTACAAGAAAA TGTGCACGCGACAGTTCTAATCGAAGAGACGATGAAGAAGCTGAAATCTGTTGTCTACGATGTGGGA AAAATTCGGGCTGATCCTATTGTCAATAGAGCTCAAATGGAGAGATGGTGGAGAAATCAAAGCACAG CGGTACAGGCTAAGGTAGTAGCAGATGTGAGAGAGTTACATGAAATAGACTATTCGTCTTACATGTA TATGATCAAATCTGACGTGAAACCTAAGACTGATTTAACACCGCAATTTGAATACTCAGCTCTACAG ACTGTTGTGTATCACGAGAAGTTGATCAACTCGTTGTTCGGTCCAATTTTCAAAGAAATTAATGAAC GCAAGTTGGATGCTATGCAACCACATTTTGTGTTCAACACGAGAATGACATCGAGTGATTTAAACGA TCGAGTGAAGTTCTTAAATACGGAAGCGGCTTACGACTTTGTTGAGATAGACATGTCTAAATTCGAC AAGTCGGCAAATCGCTTCCATTTACAACTGCAGCTGGAGATTTACAGGTTATTTGGGCTGGATGAGT GGGCGGCCTTCCTTTGGGAGGTGTCGCACACTCAAACTACTGTGAGAGATATTCAAAATGGTATGAT GGCGCATATTTGGTACCAACAAAAGAGTGGAGATGCTGATACTTATAATGCAAATTCAGATAGAACA CTGTGTGCACTCTTGTCTGAATTACCATTGGAGAAAGCAGTCATGGTTACATATGGAGGAGATGACT CACTGATTGCGTTTCCTAGAGGAACGCAGTTTGTTGATCCGTGTCCAAAGTTGGCTACTAAGTGGAA TTTCGAGTGCAAGATTTTTAAGTACGATGTCCCAATGTTTTGTGGGAAGTTCTTGCTTAAGACGTCA TCGTGTTACGAGTTCGTGCCAGATCCGGTAAAAGTTCTGACGAAGTTGGGGAAAAAGAGTATAAAGG ATGTGCAACATTTAGCCGAGATCTACATCTCGCTGAATGATTCCAATAGAGCTCTTGGGAACTACAT GGTGGTATCCAAACTGTCCGAGTCTGTTTCAGACCGGTATTTGTACAAAGGTGATTCTGTTCATGCG CTTTGTGCGCTATGGAAGCATATTAAGAGTTTTACAGCTCTGTGTACATTATTCCGAGACGAAAACG ATAAGGAATTGAACCCGGCTAAGGTTGATTGGAAGAAGGCACAGAGAGCTGTGTCAAACTTTTACGA CTGGTAATATGGAAGACAAGTCATTGGTCACCTTGAAGAAGAAGACT GGCGCGCCACGTGTTAATTA A CTGATTCGACTAGGCGCCTCAATGTGGAAGAACTGAACAGTTCGGATTACATTGAAGGCGATTTTA CCGATCAAGAGGTTTTCGGTGAGTTCATGTCTTTGAAACAAGTGGAGATGAAGACGATTGAGGCGAA GTACGATGGTCCTTACAGACCAGCTACTACTAGACCTAAGTCATTATTGTCAAGTGAAGATGTTAAG AGAGCGTCTAATAAGAAAAACTCGTCTTAATGCATAAAGAAATTTATTGTCAATATGACGTGTGTAC TCAAGGGTTGTGTGAATGAAGT...GTTCCTTTCGGGATTGATCGTTTGTTTGCTTTTTGATTTTAT TTTATATTGTTATCTGTTTCTGTGTATAGACTGTTTGAGATTGGCGCTTGGCCGACTCATTGTCTTA CCATAGGGGAACGGACTTTGTTTGTGTTGTTATTTTATTTGTATTTTATTAAAATTCTCAATGATCT GAAAAGGCCTCGAGGCTAAGAGATTATTGGGGGGTGAGTAAGTACTTTTAAAGTGATGATGGTTACA AAGGCAAAAGGGGTAAAACCCCTCGCCTACGTAAGCGTTATTACGCCCG SEQ ID NO 4 - pBTAΔRepΔMP GATCCATAAAACATTTCAATCCTTTGAACGCGGTAGAACGTGCTAATTGGATTTTGGTGAGAACGCG GTAGAACGTACTTATCACCTACAGTTTTATTTTGTTTTTCTTTTTGGTTTAATCTATCCAGCTTAGT ACCGAGTGGGGGAAAGTGACTGGTGTGCCTAAAACCTTTTCTTTGATACTTTGTAAAAATACATACA GATACAATGGCGAACGGTAACTTCAAGTTGTCTCAATTGCTCAATGTGGACGAGATGTCTGCTGAGC AGAGGAGTCATTTCTTTGACTTGATGCTGACTAAACCTGATTGTGAGATCGGGCAAATGATGCAAAG AGTTGTTGTTGATAAAGTCGATGACATGATTAGAGAAAGAAAGACTAAAGATCCAGTGATTGTTCAT GAAGTTCTTTCTCAGAAGGAACAGAACAAGTTGATGGAAATTTATCCTGAATTCAATATCGTGTTTA AAGACGACAAAAACATGGTTCATGGGTTTGCGGCTGCTGAGCGAAAACTACAAGCTTTATTGCTTTT AGATAGAGTTCCTGCTCTGCAAGAGGTGGATGACATCGGTGGTCAATGGTCGTTTTGGGTAACTAGA GGTGAGAAAAGGATTCATTCCTGTTGTCCAAATCTAGATATTCGGGATGATCAGAGAGAAATTTCTC GACAGATATTTCTTACTGCTATTGGTGATCAAGCTAGAAGTGGTAAGAGACAGATGTCGGAGAATGA GCTGTGGATGTATGACCAATTTCGTGAAAATATTGCTGCGCCTAACGCGGTTAGGTGCAATAATACA TATCAGGGTTGTACATGTAGGGGTTTTTCTGATGGTAAGAAGAAAGGCGCGCAGTATGCGATAGCTC TTCACAGCCTGTATGACTTCAAGTTGAAAGACTTGATGGCTACTATGGTTGAGAAGAAAACTAAAGT GGTTCATGCTGCTATGCTTTTTGCTCCTGAAAGTATGTTAGTGGACGAAGGTCCATTACCTTCTGTT GACGGTTACTACATGAAGAAGAACGGGAAGATCTATTTCGGTTTTGAGAAAGATCCTTCCTTTTCTT ACATTCATGACTGGGAAGAGTACAAGAAGTATCTACTGGGGAAGCCAGTGAGTTACCAAGGGAATGT GTTCTACTTCGAACCGTGGCAGGTGAGAGGAGACACAATGCTTTTTTCGATCTACAGGATAGCTGGA GTTCCGAGGAGGTCTCTATCATCGCAAGAGTACTACCGAAGAATATATATCAGTAGATGGGAAAGCA TGGTTGTTGTCCCAATTTTCGATCTGGTCGAATCAACGCGAGAGTTGGTCAAGAAAGACCTGTTTGT AGAGAAACAATTCATGGACAAGTGTTTGGATTACATAGCTAGGTTATCTGACCAGCAGCTGACCATA AGCAATGTTAAATCATACTTGAGTTCAAATAATTGGGTCTTATTCATAAACGGGGCGGCCGTGAAGA ACAAGCAAAGTGTAGATTCTCGAGATTTACAGTTGTTGGCTCAAACTTTGCTAGTGAAGGAACAAGT GGCGAGACCTGTCATGAGGGAGTTGCGTGAAGCAATTCTGACTGAGACGAAACCTATCACGTCATTG ACTGATGTGCTGGGTTTAATATCAAGAAAACTGTGGAAGCAGTTTGCTAACAAGATCGCAGTCGGCG GATTCGTTGGCATGGTTGGTACTCTAATTGGATTCTATCCAAAGAAGGTACTAACCTGGGCGAAGGA CACACCAAATGGTCCAGAACTATGTTACGAGAACTCGCACAAAACCAAGGTGATAGTATTTCTGAGT GTTGTGTATGCCATTGGAGGAATCACGCTTATGCGTCGAGACATCCGAGATGGACTGGTGAAAAAAC TATGTGATATGTTTGATATCAAACGGGGGGCCCATGTCTTAGACGTTGAGAATCCGTGCCGCTATTA TGAAATCAACGATTTCTTTAGCAGTCTGTATTCGGCATCTGAGTCCGGTGAGACCGTTTTACCAGAT TTATCCGAGGTAAAAGCCAAGTCTGATAAGCTATTGCAGCAGAAGAAAGAAATCGCTGACGAGTTTC TAAGTGCAAAATTCTCTAACTATTCTGGCAGTTCGGTGAGAACTTCTCCACCATCGGTGGTCGGTTC ATCTCGAAGCGGACTGGGTCTGTTGTTGGAAGACAGTAACGTGCTGACCCAAGCTAGAGTTGGAGTT TCAAGAAAGGTAGACGATGAGGAGATCATGGAGCAGTTTCTGAGTGGTCTTATTGACACTGAAGCAG AAATTGACGAGGTTGTTTCAGCCTTTTCAGCTGAATGTGAAAGAGGGGAAACAAGCGGTACAAAGGT GTTGTGTAAACCTTTAACGCCACCAGGATTTGAGAACGTGTTGCCAGCTGTCAAACCTTTGGTCAGC AAAGGAAAAACGGTCAAACGTGTCGATTACTTCCAAGTGATGGGAGGTGAGAGATTACCAAAAAGGC CGGTTGTCAGTGGAGACGATTCTGTGGACGCTAGAAGAGAGTTTCTGTACTACTTAGATGCGGAGAG AGTCGCTCAAAATGATGAAATTATGTCTCTGTATCGTGACTATTCGAGAGGAGTTATTCGAACTGGA GGTCAGAATTACCCGCACGGACTGGGAGTGTGGGATGTGGAGATGAAGAACTGGTGCATACGTCCAG TGGTCACTGAACATGCTTATGTGTTCCAACCAGACAAACGTATGGATGATTGGTCGGGATACTTAGA AGTGGCTGTTTGGGAACGAGGTATGTTGGTCAACGACTTCGCGGTCGAAAGGATGAGTGATTATGTC ATAGTTTGCGATCAGACGTATCTTTGCAATAACAGGTAATAATCCTCTCTCTTGATATTTTTAAATT ATAGAATTAATTAGTTTACTTTATTCTTTACTATATGATTT(...)AACTGTTCTTTTGAATTAGTTG ACGGTGTACCTGGTTGTGGTAAGTCGACAATGATTGTCAACTCAGCTAATCCTTGTGTCGATGTGGT TCTCTCTACTGGGAGAGCAGCAACCGACGACTTGATCGAGAGATTCGCGAGCAAAGGTTTTCCATGC AAATTGAAAAGGAGAGTGAAGACGGTTGATTCTTTTTTGATGCATTGTGTCGATGGTTCTTTAACCG GAGACGTGTTGCATTTCGACGAAGCTCTCATGGCCCATGCTGGTATGGTGTACTTTTGCGCTCAGAT AGCTGGTGCTAAACGATGTATCTGTCAAGGAGATCAGAATCAAATTTCTTTCAAGCCTAGGGTATCT CAAGTTGATTTGAGGTTTTCTAGTCTGGTCGGAAAGTTTGACATTGTTACAGAAAAAAGAGAAACTT ACAGAAGTCCAGCAGATGTGGCTGCCGTATTGAACAAGTACTATACTGGAGATGTCAGAACACATAA CGCGACTGCTAATTCGATGACGGTGAGGAAGATTGTGTCTAAAGAACAGGTTTCTTTGAAGCCTGGT GCTCAGTACATAACTTTCCTTCAGTCTGAGAAGAAGGAGTTGGTAAATTTGTTGGCATTGAGGAAAG TGGCAGCTAAAGTGAGTACAGTACACGAGTCGCAAGGAGAGACATTCAAAGATGTAGTCCTAGTCAG GACGAAACCTACGGATGACTCAATCGCTAGAGGTCGGGAGTACTTAATCGTGGCATTGTCGCGTCAC ACACAATCACTTGTGTATGAAACTGTGAAAGAGGACGATGTAAGCAAAGAGATCAGGGAAAGTGCCG CGCTTACGAAGGCGGCTTTGGCAAGATTTTTTGTTACTGAGACCGTCTTATGACGGTTTCGGTCTAG GTTTGATGTCTTTAGACATCATGAAGGGCCTTGCGCCGTTCCAGATTCAGGTACGATTACGGACTTG GAGATGTGGTACGACGCTTTGTTTCCGGGAAATTCGTTAAGAGACTCAAGCCTAGACGGGTATTTGG TGGCAACGACTGATTGCAATTTGCGATTAGACAATGTTACGATCAAAAGTGGAAACTGGAAAGACAA GTTTGCTGAAAAAGAAACGTTTCTGAAACCGGTTATTCGTACTGCTATGCCTGACAAAAGGAAGACT ACTCAGTTGGAGAGTTTGTTAGCATTGCAGAAAAGGAACCAAGCGGCACCCGATCTACAAGAAAATG TGCACGCGACAGTTCTAATCGAAGAGACGATGAAGAAGCTGAAATCTGTTGTCTACGATGTGGGAAA AATTCGGGCTGATCCTATTGTCAATAGAGCTCAAATGGAGAGATGGTGGAGAAATCAAAGCACAGCG GTACAGGCTAAGGTAGTAGCAGATGTGAGAGAGTTACATGAAATAGACTATTCGTCTTACATGTATA TGATCAAATCTGACGTGAAACCTAAGACTGATTTAACACCGCAATTTGAATACTCAGCTCTACAGAC TGTTGTGTATCACGAGAAGTTGATCAACTCGTTGTTCGGTCCAATTTTCAAAGAAATTAATGAACGC AAGTTGGATGCTATGCAACCACATTTTGTGTTCAACACGAGAATGACATCGAGTGATTTAAACGATC GAGTGAAGTTCTTAAATACGGAAGCGGCTTACGACTTTGTTGAGATAGACATGTCTAAATTCGACAA GTCGGCAAATCGCTTCCATTTACAACTGCAGCTGGAGATTTACAGGTTATTTGGGCTGGATGAGTGG GCGGCCTTCCTTTGGGAGGTGTCGCACACTCAAACTACTGTGAGAGATATTCAAAATGGTATGATGG CGCATATTTGGTACCAACAAAAGAGTGGAGATGCTGATACTTATAATGCAAATTCAGATAGAACACT GTGTGCACTCTTGTCTGAATTACCATTGGAGAAAGCAGTCATGGTTACATATGGAGGAGATGACTCA CTGATTGCGTTTCCTAGAGGAACGCAGTTTGTTGATCCGTGTCCAAAGTTGGCTACTAAGTGGAATT TCGAGTGCAAGATTTTTAAGTACGATGTCCCAATGTTTTGTGGGAAGTTCTTGCTTAAGACGTCATC GTGTTACGAGTTCGTGCCAGATCCGGTAAAAGTTCTGACGAAGTTGGGGAAAAAGAGTATAAAGGAT GTGCAACATTTAGCCGAGATCTACATCTCGCTGAATGATTCCAATAGAGCTCTTGGGAACTACATGG TGGTATCCAAACTGTCCGAGTCTGTTTCAGACCGGTATTTGTACAAAGGTGATTCTGTTCATGCGCT TTGTGCGCTATGGAAGCATATTAAGAGTTTTACAGCTCTGTGTACATTATTCCGAGACGAAAACGAT AAGGAATTGAACCCGGCTAAGGTTGATTGGAAGAAGGCACAGAGAGCTGTGTCAAACTTTTACGACT GGTAATATGGAAGACAAGTCATTGGTCACCTTGAAGAAGAAGACT GGCGCGCCACGTGTTAATTAA C TGATTCGACTAGGCGCCTCAATGTGGAAGAACTGAACAGTTCGGATTACATTGAAGGCGATTTTACC GATCAAGAGGTTTTCGGTGAGTTCATGTCTTTGAAACAAGTGGAGATGAAGACGATTGAGGCGAAGT ACGATGGTCCTTACAGACCAGCTACTACTAGACCTAAGTCATTATTGTCAAGTGAAGATGTTAAGAG AGCGTCTAATAAGAAAAACTCGTCTTAATGCATAAAGAAATTTATTGTCAATATGACGTGTGTACTC AAGGGTTGTGTGAATGAAGTCACTGTTCTTGGTCACGAGACGTGTAGTATCGGTCATGCTAACAAAT TGCGAAAGCAAGTTGCTGACATGGTTGGTGTCACACGTAGGTGTGCGGAAAATAATTGTGGATGGTT TGTCTGTGTTGTTATCAATGATTTTACTTTTGATGTGTATAATTGTTGTGGCCGTAGTCACCTTGAA AAGTGTCGTAAACGTGTTGAAACAAGAAATCGAGAAATTTGGAAACAAATTCGACGAAATCAAGCTG AAAACATGTCTGCGACAGCTAAAAAGTCTCATAATTCGAAGACCTCTAAGAAGAAATTCAAAGAGGA CAGAGAATTTGGGACACCAAAAAGATTTTTAAGAGATGATGTTCCTTTCGGGATTGATCGTTTGTTT GCTTTTTGATTTTATTTTATATTGTTATCTGTTTCTGTGTATAGACTGTTTGAGATTGGCGCTTGGC CGACTCATTGTCTTACCATAGGGGAACGGACTTTGTTTGTGTTGTTATTTTATTTGTATTTTATTAA AATTCTCAATGATCTGAAAAGGCCTCGAGGCTAAGAGATTATTGGGGGGTGAGTAAGTACTTTTAAA GTGATGATGGTTACAAAGGCAAAAGGGGTAAAACCCCTCGCCTACGTAAGCGTTATTACGCCCG SEQ ID NO 5 - pBTAΔRepΔM A16K GATCCATAAAACATTTCAATCCTTTGAACGCGGTAGAACGTGCTAATTGGATTTTGGTGAGAACGCG GTAGAACGTACTTATCACCTACAGTTTTATTTTGTTTTTCTTTTTGGTTTAATCTATCCAGCTTAGT ACCGAGTGGGGGAAAGTGACTGGTGTGCCTAAAACCTTTTCTTTGATACTTTGTAAAAATACATACA GATACAATGGCGAACGGTAACTTCAAGTTGTCTCAATTGCTCAATGTGGACGAGATGTCTGCTGAGC AGAGGAGTCATTTCTTTGACTTGATGCTGACTAAACCTGATTGTGAGATCGGGCAAATGATGCAAAG AGTTGTTGTTGATAAAGTCGATGACATGATTAGAGAAAGAAAGACTAAAGATCCAGTGATTGTTCAT GAAGTTCTTTCTCAGAAGGAACAGAACAAGTTGATGGAAATTTATCCTGAATTCAATATCGTGTTTA AAGACGACAAAAACATGGTTCATGGGTTTGCGGCTGCTGAGCGAAAACTACAAGCTTTATTGCTTTT AGATAGAGTTCCTGCTCTGCAAGAGGTGGATGACATCGGTGGTCAATGGTCGTTTTGGGTAACTAGA GGTGAGAAAAGGATTCATTCCTGTTGTCCAAATCTAGATATTCGGGATGATCAGAGAGAAATTTCTC GACAGATATTTCTTACTGCTATTGGTGATCAAGCTAGAAGTGGTAAGAGACAGATGTCGGAGAATGA GCTGTGGATGTATGACCAATTTCGTGAAAATATTGCTGCGCCTAACGCGGTTAGGTGCAATAATACA TATCAGGGTTGTACATGTAGGGGTTTTTCTGATGGTAAGAAGAAAGGCGCGCAGTATGCGATAGCTC TTCACAGCCTGTATGACTTCAAGTTGAAAGACTTGATGGCTACTATGGTTGAGAAGAAAACTAAAGT GGTTCATGCTGCTATGCTTTTTGCTCCTGAAAGTATGTTAGTGGACGAAGGTCCATTACCTTCTGTT GACGGTTACTACATGAAGAAGAACGGGAAGATCTATTTCGGTTTTGAGAAAGATCCTTCCTTTTCTT ACATTCATGACTGGGAAGAGTACAAGAAGTATCTACTGGGGAAGCCAGTGAGTTACCAAGGGAATGT GTTCTACTTCGAACCGTGGCAGGTGAGAGGAGACACAATGCTTTTTTCGATCTACAGGATAGCTGGA GTTCCGAGGAGGTCTCTATCATCGCAAGAGTACTACCGAAGAATATATATCAGTAGATGGGAAAGCA TGGTTGTTGTCCCAATTTTCGATCTGGTCGAATCAACGCGAGAGTTGGTCAAGAAAGACCTGTTTGT AGAGAAACAATTCATGGACAAGTGTTTGGATTACATAGCTAGGTTATCTGACCAGCAGCTGACCATA AGCAATGTTAAATCATACTTGAGTTCAAATAATTGGGTCTTATTCATAAACGGGGCGGCCGTGAAGA ACAAGCAAAGTGTAGATTCTCGAGATTTACAGTTGTTGGCTCAAACTTTGCTAGTGAAGGAACAAGT GGCGAGACCTGTCATGAGGGAGTTGCGTGAAGCAATTCTGACTGAGACGAAACCTATCACGTCATTG ACTGATGTGCTGGGTTTAATATCAAGAAAACTGTGGAAGCAGTTTGCTAACAAGATCGCAGTCGGCG GATTCGTTGGCATGGTTGGTACTCTAATTGGATTCTATCCAAAGAAGGTACTAACCTGGGCGAAGGA CACACCAAATGGTCCAGAACTATGTTACGAGAACTCGCACAAAACCAAGGTGATAGTATTTCTGAGT GTTGTGTATGCCATTGGAGGAATCACGCTTATGCGTCGAGACATCCGAGATGGACTGGTGAAAAAAC TATGTGATATGTTTGATATCAAACGGGGGGCCCATGTCTTAGACGTTGAGAATCCGTGCCGCTATTA TGAAATCAACGATTTCTTTAGCAGTCTGTATTCGGCATCTGAGTCCGGTGAGACCGTTTTACCAGAT TTATCCGAGGTAAAAGCCAAGTCTGATAAGCTATTGCAGCAGAAGAAAGAAATCGCTGACGAGTTTC TAAGTGCAAAATTCTCTAACTATTCTGGCAGTTCGGTGAGAACTTCTCCACCATCGGTGGTCGGTTC ATCTCGAAGCGGACTGGGTCTGTTGTTGGAAGACAGTAACGTGCTGACCCAAGCTAGAGTTGGAGTT TCAAGAAAGGTAGACGATGAGGAGATCATGGAGCAGTTTCTGAGTGGTCTTATTGACACTGAAGCAG AAATTGACGAGGTTGTTTCAGCCTTTTCAGCTGAATGTGAAAGAGGGGAAACAAGCGGTACAAAGGT GTTGTGTAAACCTTTAACGCCACCAGGATTTGAGAACGTGTTGCCAGCTGTCAAACCTTTGGTCAGC AAAGGAAAAACGGTCAAACGTGTCGATTACTTCCAAGTGATGGGAGGTGAGAGATTACCAAAAAGGC CGGTTGTCAGTGGAGACGATTCTGTGGACGCTAGAAGAGAGTTTCTGTACTACTTAGATGCGGAGAG AGTCGCTCAAAATGATGAAATTATGTCTCTGTATCGTGACTATTCGAGAGGAGTTATTCGAACTGGA GGTCAGAATTACCCGCACGGACTGGGAGTGTGGGATGTGGAGATGAAGAACTGGTGCATACGTCCAG TGGTCACTGAACATGCTTATGTGTTCCAACCAGACAAACGTATGGATGATTGGTCGGGATACTTAGA AGTGGCTGTTTGGGAACGAGGTATGTTGGTCAACGACTTCGCGGTCGAAAGGATGAGTGATTATGTC ATAGTTTGCGATCAGACGTATCTTTGCAATAACAGGTAATAATCCTCTCTCTTGATATTTTTAAATT ATAGAATTAATTAGTTTACTTTATTCTTTACTATATGATTT(...)AACTGTTCTTTTGAATTAGTTG ACGGTGTACCTGGTTGTGGTAAGTCGACAATGATTGTCAACTCAGCTAATCCTTGTGTCGATGTGGT TCTCTCTACTGGGAGAGCAGCAACCGACGACTTGATCGAGAGATTCGCGAGCAAAGGTTTTCCATGC AAATTGAAAAGGAGAGTGAAGACGGTTGATTCTTTTTTGATGCATTGTGTCGATGGTTCTTTAACCG GAGACGTGTTGCATTTCGACGAAGCTCTCATGGCCCATGCTGGTATGGTGTACTTTTGCGCTCAGAT AGCTGGTGCTAAACGATGTATCTGTCAAGGAGATCAGAATCAAATTTCTTTCAAGCCTAGGGTATCT CAAGTTGATTTGAGGTTTTCTAGTCTGGTCGGAAAGTTTGACATTGTTACAGAAAAAAGAGAAACTT ACAGAAGTCCAGCAGATGTGGCTGCCGTATTGAACAAGTACTATACTGGAGATGTCAGAACACATAA CGCGACTGCTAATTCGATGACGGTGAGGAAGATTGTGTCTAAAGAACAGGTTTCTTTGAAGCCTGGT GCTCAGTACATAACTTTCCTTCAGTCTGAGAAGAAGGAGTTGGTAAATTTGTTGGCATTGAGGAAAG TGGCAGCTAAAGTGAGTACAGTACACGAGTCGCAAGGAGAGACATTCAAAGATGTAGTCCTAGTCAG GACGAAACCTACGGATGACTCAATCGCTAGAGGTCGGGAGTACTTAATCGTGGCATTGTCGCGTCAC ACACAATCACTTGTGTATGAAACTGTGAAAGAGGACGATGTAAGCAAAGAGATCAGGGAAAGTGCCG CGCTTACGAAGGCGGCTTTGGCAAGATTTTTTGTTACTGAGACCGTCTTATGACGGTTTCGGTCTAG GTTTGATGTCTTTAGACATCATGAAGGGCCTTGCGCCGTTCCAGATTCAGGTACGATTACGGACTTG GAGATGTGGTACGACGCTTTGTTTCCGGGAAATTCGTTAAGAGACTCAAGCCTAGACGGGTATTTGG TGGCAACGACTGATTGCAATTTGCGATTAGACAATGTTACGATCAAAAGTGGAAACTGGAAAGACAA GTTTGCTGAAAAAGAAACGTTTCTGAAACCGGTTATTCGTACTGCTATGCCTGACAAAAGGAAGACT ACTCAGTTGGAGAGTTTGTTAGCATTGCAGAAAAGGAACCAAGCGGCACCCGATCTACAAGAAAATG TGCACGCGACAGTTCTAATCGAAGAGACGATGAAGAAGCTGAAATCTGTTGTCTACGATGTGGGAAA AATTCGGGCTGATCCTATTGTCAATAGAGCTCAAATGGAGAGATGGTGGAGAAATCAAAGCACAGCG GTACAGGCTAAGGTAGTAGCAGATGTGAGAGAGTTACATGAAATAGACTATTCGTCTTACATGTATA TGATCAAATCTGACGTGAAACCTAAGACTGATTTAACACCGCAATTTGAATACTCAGCTCTACAGAC TGTTGTGTATCACGAGAAGTTGATCAACTCGTTGTTCGGTCCAATTTTCAAAGAAATTAATGAACGC AAGTTGGATGCTATGCAACCACATTTTGTGTTCAACACGAGAATGACATCGAGTGATTTAAACGATC GAGTGAAGTTCTTAAATACGGAAGCGGCTTACGACTTTGTTGAGATAGACATGTCTAAATTCGACAA GTCGGCAAATCGCTTCCATTTACAACTGCAGCTGGAGATTTACAGGTTATTTGGGCTGGATGAGTGG GCGGCCTTCCTTTGGGAGGTGTCGCACACTCAAACTACTGTGAGAGATATTCAAAATGGTATGATGG CGCATATTTGGTACCAACAAAAGAGTGGAGATGCTGATACTTATAATGCAAATTCAGATAGAACACT GTGTGCACTCTTGTCTGAATTACCATTGGAGAAAGCAGTCATGGTTACATATGGAGGAGATGACTCA CTGATTGCGTTTCCTAGAGGAACGCAGTTTGTTGATCCGTGTCCAAAGTTGGCTACTAAGTGGAATT TCGAGTGCAAGATTTTTAAGTACGATGTCCCAATGTTTTGTGGGAAGTTCTTGCTTAAGACGTCATC GTGTTACGAGTTCGTGCCAGATCCGGTAAAAGTTCTGACGAAGTTGGGGAAAAAGAGTATAAAGGAT GTGCAACATTTAGCCGAGATCTACATCTCGCTGAATGATTCCAATAGAGCTCTTGGGAACTACATGG TGGTATCCAAACTGTCCGAGTCTGTTTCAGACCGGTATTTGTACAAAGGTGATTCTGTTCATGCGCT TTGTGCGCTATGGAAGCATATTAAGAGTTTTACAGCTCTGTGTACATTATTCCGAGACGAAAACGAT AAGGAATTGAACCCGGCTAAGGTTGATTGGAAGAAGGCACAGAGAGCTGTGTCAAACTTTTACGACT GGTAATATGGAAGACAAGTCATTGGTCACCTTGAAGAAGAAGACT GGCGCGCCACGTGTTAATTAA C TGATTCGACTAGGCGCCTCAATGTGGAAGAACTGAACAGTTCGGATTACATTGAAGGCGATTTTACC GATCAAGAGGTTTTCGGTGAGTTCATGTCTTTGAAACAAGTGGAGATGAAGACGATTGAGGCGAAGT ACGATGGTCCTTACAGACCAGCTACTACTAGACCTAAGTCATTATTGTCAAGTGAAGATGTTAAGAG AGCGTCTAATAAGAAAAACTCGTCTTAATGCATAAAGAAATTTATTGTCAATATGACGTGTGTACTC AAGGGTTGTGTGAATGAAGT...GTTCCTTTCGGGATTGATCGTTTGTTTGCTTTTTGATTTTATTT TATATTGTTATCTGTTTCTGTGTATAGACTGTTTGAGATTGGCGCTTGGCCGACTCATTGTCTTACC ATAGGGGAACGGACTTTGTTTGTGTTGTTATTTTATTTGTATTTTATTAAAATTCTCAATGATCTGA AAAGGCCTCGAGGCTAAGAGATTATTGGGGGGTGAGTAAGTACTTTTAAAGTGATGATGGTTACAAA GGCAAAAGGGGTAAAACCCCTCGCCTACGTAAGCGTTATTACGCCCG

Sequences Inserted Into the Amplicon Constructs

[0171] SEQ ID NO 6 - A. thaliana Partial CDNA Sequence Sulphur Gene. ccttcactctcttctccttcctcaaaaccttcctcctcccccatttgcttcaggccaggtaaattgt ttggaagcaagttaaatgcaggaatccaaataaggccaaagaagaacaggtctcgttaccatgtttc ggttatgaatgtagccactgaaatcaactctactgaacaagtagtagggaagtttgattcaaagaag agtgcgagaccggtttatccatttgcagctatagtagggcaagatgagatgaagttatgtcttttgt tgaatgttattgatccaaagattggtggtgttatgattatgggagatagaggaactggaaaatctac aactgttagatcattagttgatctgttacctgagattaatgtagttgcaggtgacccgtataactcg gatccgatagatcctgagtttatgggtgttgaagtaagagagagagttgagaaaggagagcaagttc ctgttattgcgactaagattaatatggttgatcttcctttgggtgcaacagaagatagagtttgtgg aaccatcgatatcgaaaaggctttgacagaaggtgtaaaagcctttgagcctggtttgttggctaaa gctaatagagggattctttatgttgatgaagttaatctcttggatgatcatttggttgatgttcttt tggattcagctgcttctggttggaatacggttgagagagaagggatttcgatttctcacccggcgag gtttatcttgatcggttcaggaaatccggaagaaggagagcttaggccacagcttcttgatcggttt ggtatgcatgcacaagtagggacggttagagatgctgatttacgggtcaagattgttgaagagagag ctcgtttcgatagtaacccaaaggatttccgtgacacttacaaaaccgagcaggacaagcttcaaga ccagatt SEQ ID NO 7 - A. thaliana Partial cDNA Sequence RUBISCO Small Sub-Unit Gene. cctctatgctctcttccgctactatggttgcctctccggctcaggccactatggtcgctcctttcaa cggacttaagtcctccgctgccttcccagccacccgcaaggctaacaacgacattacttccatcaca agcaacggcggaagagttaactgcatgcaggtgtggcctccgattggaaagaagaagtttgagactc tctcttaccttcctgaccttaccgattccgaattggctaaggaagttgactaccttatccgcaacaa gtggattccttgtgttgaattcgagttggagcacggatttgtgtaccgtgagcacggtaactcaccc ggatactatgatggacggtactggacaatgtggaagcttcccttgttcggttgcaccgactccgctc aagtgttgaaggaagtggaagagtgcaagaaggagtaccccaatgccttcattaggatcatcggatt SEQ ID NO 8 - A. thaliana Partial cDNA Sequence LEAFY Gene. ccatacggtatacgtttctacacggcggcgaagatagcggagttaggttttacggcgagcacgcttg tgggtatgaaggacgaggagcttgaagagatgatgaatagtctctctcatatctttcgttgggagct tcttgttggtgaacggtacggtatcaaagctgccgttagagctgaacggagacgattgcaagaagag gaggaagaggaatcttctagacgccgtcatttgctactctccgccgctggtgattccggtactcatc acgctcttgatgctctctcccaagaagtgattggacagggttatctgaggaaccggtgcagcaacaa gaccagactgatgcggcggggaataacggcggaggaggaagtggttactgggacgcaggtcaaggaa agatgaagaagcaacagcagcagagacggagaaagaaaccaatgctgacgtcagtggaaaccgacga agacgtcaacgaaggtgaggatgacgacgggatggataacggcaacggaggtagtggtttggggaca gagagacagagggagcatccgtttatcgtaacggagcctggggaagtggcacgtggcaaaaagaacg gcttagattatctgttccacttgtacgaacaatgccgtgagttccttcttcaggtccagacaattgc taaagaccgtggcgaaaaatgccccaccaaggtgacgaaccaagtattcaggtacgcgaagaaatca ggagcgagttacataaacaagcctaaaatgcgacactacgttcactgttacgctctccactgcctag acgaagaagcttcaaatgctctcagaagagcgtttaaagaacgcggtgagaacgttggctcatggcg tcaggcttgttacaagccacttgtgaacatcgcttgtcgtcatggctgggatatagacgccgtcttt aa SEQ ID NO 9 - mGFP5 cDNA Sequence. aatgaagactaatctttttctctttctcatcttttcacttctcctatcattatcctcggccgaattc agtaaaggagaagaacttttcactggagttgtcccaattcttgttgaattagatggtgatgttaatg ggcacaaattttctgtcagtggagagggtgaaggtgatgcaacatacggaaaacttacccttaaatt tatttgcactactggaaaactacctgttccatggccaacacttgtcactactttctcttatggtgtt caatgcttttcaagatacccagatcatatgaagcggcacgacttcttcaagagcgccatgcctgagg gatacgtgcaggagaggaccatcttcttcaaggacgacgggaactacaagacacgtgctgaagtcaa gtttgagggagacaccctcgtcaacaggatcgagcttaagggaatcgatttcaaggaggacggaaac atcctcggccacaagttggaatacaactacaactcccacaacgtatacatcatggccgacaagcaga agaacggcatcaaagccaacttcaagacccgccacaacatcgaagacggcggcgtgcaactcgctga tcattatcaacaaaatactccaattggcgatggccctgtccttttaccagacaaccattacctgtcc acacaatctgccctttcgaaagatcccaacgaaaagagagaccacatggtccttcttgagtttgtaa cagctgctgggattacacatggcatggatgaactatacaaacatgacgaactctaa

[0172]

1 32 1 20174 DNA Artificial Sequence Description of Artificial Sequence pBINTRA6 1 tactccaaaa atgtcaaaga tacagtctca gaagaccaaa gggctattga gacttttcaa 60 caaagggtaa tttcgggaaa cctcctcgga ttccattgcc cagctatctg tcacttcatc 120 gaaaggacag tagaaaagga aggtggctcc tacaaatgcc atcattgcga taaaggaaag 180 gctatcattc aagatgcctc tgccgacagt ggtcccaaag atggaccccc acccacgagg 240 agcatcgtgg aaaaagaaga cgtcccaacc acgtcttcaa agcaagtgga ttgatgtgac 300 atctccactg acgtaaggga tgacgcacaa tcccactatc cttcgcaaga cccttcttct 360 atataaggaa gttcatttca tttggagagg acagcccaag ctttctagag gatccataaa 420 acatttcaat cctttgaacg cggtagaacg tgctaattgg attttggtga gaacgcggta 480 gaacgtactt atcacctaca gttttatttt gtttttcttt ttggtttaat ctatccagct 540 tagtaccgag tgggggaaag tgactggtgt gcctaaaacc ttttctttga tactttgtaa 600 aaatacatac agatacaatg gcgaacggta acttcaagtt gtctcaattg ctcaatgtgg 660 acgagatgtc tgctgagcag aggagtcatt tctttgactt gatgctgact aaacctgatt 720 gtgagatcgg gcaaatgatg caaagagttg ttgttgataa agtcgatgac atgattagag 780 aaagaaagac taaagatcca gtgattgttc atgaagttct ttctcagaag gaacagaaca 840 agttgatgga aatttatcct gaattcaata tcgtgtttaa agacgacaaa aacatggttc 900 atgggtttgc ggctgctgag cgaaaactac aagctttatt gcttttagat agagttcctg 960 ctctgcaaga ggtggatgac atcggtggtc aatggtcgtt ttgggtaact agaggtgaga 1020 aaaggattca ttcctgttgt ccaaatctag atattcggga tgatcagaga gaaatttctc 1080 gacagatatt tcttactgct attggtgatc aagctagaag tggtaagaga cagatgtcgg 1140 agaatgagct gtggatgtat gaccaatttc gtgaaaatat tgctgcgcct aacgcggtta 1200 ggtgcaataa tacatatcag ggttgtacat gtaggggttt ttctgatggt aagaagaaag 1260 gcgcgcagta tgcgatagct cttcacagcc tgtatgactt caagttgaaa gacttgatgg 1320 ctactatggt tgagaagaaa actaaagtgg ttcatgctgc tatgcttttt gctcctgaaa 1380 gtatgttagt ggacgaaggt ccattacctt ctgttgacgg ttactacatg aagaagaacg 1440 ggaagatcta tttcggtttt gagaaagatc cttccttttc ttacattcat gactgggaag 1500 agtacaagaa gtatctactg gggaagccag tgagttacca agggaatgtg ttctacttcg 1560 aaccgtggca ggtgagagga gacacaatgc ttttttcgat ctacaggata gctggagttc 1620 cgaggaggtc tctatcatcg caagagtact accgaagaat atatatcagt agatgggaaa 1680 gcatggttgt tgtcccaatt ttcgatctgg tcgaatcaac gcgagagttg gtcaagaaag 1740 acctgtttgt agagaaacaa ttcatggaca agtgtttgga ttacatagct aggttatctg 1800 accagcagct gaccataagc aatgttaaat catacttgag ttcaaataat tgggtcttat 1860 tcataaacgg ggcggccgtg aagaacaagc aaagtgtaga ttctcgagat ttacagttgt 1920 tggctcaaac tttgctagtg aaggaacaag tggcgagacc tgtcatgagg gagttgcgtg 1980 aagcaattct gactgagacg aaacctatca cgtcattgac tgatgtgctg ggtttaatat 2040 caagaaaact gtggaagcag tttgctaaca agatcgcagt cggcggattc gttggcatgg 2100 ttggtactct aattggattc tatccaaaga aggtactaac ctgggcgaag gacacaccaa 2160 atggtccaga actatgttac gagaactcgc acaaaaccaa ggtgatagta tttctgagtg 2220 ttgtgtatgc cattggagga atcacgctta tgcgtcgaga catccgagat ggactggtga 2280 aaaaactatg tgatatgttt gatatcaaac ggggggccca tgtcttagac gttgagaatc 2340 cgtgccgcta ttatgaaatc aacgatttct ttagcagtct gtattcggca tctgagtccg 2400 gtgagaccgt tttaccagat ttatccgagg taaaagccaa gtctgataag ctattgcagc 2460 agaagaaaga aatcgctgac gagtttctaa gtgcaaaatt ctctaactat tctggcagtt 2520 cggtgagaac ttctccacca tcggtggtcg gttcatctcg aagcggactg ggtctgttgt 2580 tggaagacag taacgtgctg acccaagcta gagttggagt ttcaagaaag gtagacgatg 2640 aggagatcat ggagcagttt ctgagtggtc ttattgacac tgaagcagaa attgacgagg 2700 ttgtttcagc cttttcagct gaatgtgaaa gaggggaaac aagcggtaca aaggtgttgt 2760 gtaaaccttt aacgccacca ggatttgaga acgtgttgcc agctgtcaaa cctttggtca 2820 gcaaaggaaa aacggtcaaa cgtgtcgatt acttccaagt gatgggaggt gagagattac 2880 caaaaaggcc ggttgtcagt ggagacgatt ctgtggacgc tagaagagag tttctgtact 2940 acttagatgc ggagagagtc gctcaaaatg atgaaattat gtctctgtat cgtgactatt 3000 cgagaggagt tattcgaact ggaggtcaga attacccgca cggactggga gtgtgggatg 3060 tggagatgaa gaactggtgc atacgtccag tggtcactga acatgcttat gtgttccaac 3120 cagacaaacg tatggatgat tggtcgggat acttagaagt ggctgtttgg gaacgaggta 3180 tgttggtcaa cgacttcgcg gtcgaaagga tgagtgatta tgtcatagtt tgcgatcaga 3240 cgtatctttg caataacagg taataatcct ctctcttgat atttttaaat tatagaatta 3300 attagtttac tttattcttt actatatgat ttaaatagtt taatcttgtt tttgagtaaa 3360 ctattcgatt ttgatatttg tattcgtcct acaaagttgg aaatactgat gatattttct 3420 tttgaacgtg atacctacca atactaatct tacggaatct tttaatagag cactaatcaa 3480 catggaacta aagaccaatt cttaagtgtc tctgttgtac agttcatttt agtagtgcgt 3540 ttaagtatta ttatctccct tcatgcgggg caattatgta gattaaaatc gaaattatat 3600 aaaatttaca taagtctaag tctagggtct ccagctaatt gttatttttt taacgatgtt 3660 gactaaagca ataacgacgt tgacttgtgt taaacaggtt gatcttggac aatttaagtg 3720 ccctggatct aggaccagtt aactgttctt ttgaattagt tgacggtgta cctggttgtg 3780 gtaagtcgac aatgattgtc aactcagcta atccttgtgt cgatgtggtt ctctctactg 3840 ggagagcagc aaccgacgac ttgatcgaga gattcgcgag caaaggtttt ccatgcaaat 3900 tgaaaaggag agtgaagacg gttgattctt ttttgatgca ttgtgtcgat ggttctttaa 3960 ccggagacgt gttgcatttc gacgaagctc tcatggccca tgctggtatg gtgtactttt 4020 gcgctcagat agctggtgct aaacgatgta tctgtcaagg agatcagaat caaatttctt 4080 tcaagcctag ggtatctcaa gttgatttga ggttttctag tctggtcgga aagtttgaca 4140 ttgttacaga aaaaagagaa acttacagaa gtccagcaga tgtggctgcc gtattgaaca 4200 agtactatac tggagatgtc agaacacata acgcgactgc taattcgatg acggtgagga 4260 agattgtgtc taaagaacag gtttctttga agcctggtgc tcagtacata actttccttc 4320 agtctgagaa gaaggagttg gtaaatttgt tggcattgag gaaagtggca gctaaagtga 4380 gtacagtaca cgagtcgcaa ggagagacat tcaaagatgt agtcctagtc aggacgaaac 4440 ctacggatga ctcaatcgct agaggtcggg agtacttaat cgtggcattg tcgcgtcaca 4500 cacaatcact tgtgtatgaa actgtgaaag aggacgatgt aagcaaagag atcagggaaa 4560 gtgccgcgct tacgaaggcg gctttggcaa gattttttgt tactgagacc gtcttatgac 4620 ggtttcggtc taggtttgat gtctttagac atcatgaagg gccttgcgcc gttccagatt 4680 caggtacgat tacggacttg gagatgtggt acgacgcttt gtttccggga aattcgttaa 4740 gagactcaag cctagacggg tatttggtgg caacgactga ttgcaatttg cgattagaca 4800 atgttacgat caaaagtgga aactggaaag acaagtttgc tgaaaaagaa acgtttctga 4860 aaccggttat tcgtactgct atgcctgaca aaaggaagac tactcagttg gagagtttgt 4920 tagcattgca gaaaaggaac caagcggcac ccgatctaca agaaaatgtg cacgcgacag 4980 ttctaatcga agagacgatg aagaagctga aatctgttgt ctacgatgtg ggaaaaattc 5040 gggctgatcc tattgtcaat agagctcaaa tggagagatg gtggagaaat caaagcacag 5100 cggtacaggc taaggtagta gcagatgtga gagagttaca tgaaatagac tattcgtctt 5160 acatgtatat gatcaaatct gacgtgaaac ctaagactga tttaacaccg caatttgaat 5220 actcagctct acagactgtt gtgtatcacg agaagttgat caactcgttg ttcggtccaa 5280 ttttcaaaga aattaatgaa cgcaagttgg atgctatgca accacatttt gtgttcaaca 5340 cgagaatgac atcgagtgat ttaaacgatc gagtgaagtt cttaaatacg gaagcggctt 5400 acgactttgt tgagatagac atgtctaaat tcgacaagtc ggcaaatcgc ttccatttac 5460 aactgcagct ggagatttac aggttatttg ggctggatga gtgggcggcc ttcctttggg 5520 aggtgtcgca cactcaaact actgtgagag atattcaaaa tggtatgatg gcgcatattt 5580 ggtaccaaca aaagagtgga gatgctgata cttataatgc aaattcagat agaacactgt 5640 gtgcactctt gtctgaatta ccattggaga aagcagtcat ggttacatat ggaggagatg 5700 actcactgat tgcgtttcct agaggaacgc agtttgttga tccgtgtcca aagttggcta 5760 ctaagtggaa tttcgagtgc aagattttta agtacgatgt cccaatgttt tgtgggaagt 5820 tcttgcttaa gacgtcatcg tgttacgagt tcgtgccaga tccggtaaaa gttctgacga 5880 agttggggaa aaagagtata aaggatgtgc aacatttagc cgagatctac atctcgctga 5940 atgattccaa tagagctctt gggaactaca tggtggtatc caaactgtcc gagtctgttt 6000 cagaccggta tttgtacaaa ggtgattctg ttcatgcgct ttgtgcgcta tggaagcata 6060 ttaagagttt tacagctctg tgtacattat tccgagacga aaacgataag gaattgaacc 6120 cggctaaggt tgattggaag aaggcacaga gagctgtgtc aaacttttac gactggtaat 6180 atggaagaca agtcattggt caccttgaag aagaagactt tcgaagtctc aaaattctca 6240 aatctagggg ccattgaatt gtttgtggac ggtaggagga agagaccgaa gtattttcac 6300 agaagaagag aaactgtcct aaatcatgtt ggtgggaaga agagtgaaca caagttagac 6360 gtttttgacc aaagggatta caaaatgatt aaatcttacg cgtttctaaa ggtagtaggt 6420 gtacaactag ttgtaacatc acatctacct gcagatacgc ctgggttcat tcaaatcgat 6480 ctgttggatt cgagacttac tgagaaaaga aagagaggaa agactattca gagattcaaa 6540 gctcgagctt gcgataactg ttcagttgcg cagtacaagg ttgaatacag tatttccaca 6600 caggagaacg tacttgatgt ctggaaggtg ggttgtattt ctgagggcgt tccggtctgt 6660 gacggtacat accctttcag tatcgaagtg tcgctaatat gggttgctac tgattcgact 6720 aggcgcctca atgtggaaga actgaacagt tcggattaca ttgaaggcga ttttaccgat 6780 caagaggttt tcggtgagtt catgtctttg aaacaagtgg agatgaagac gattgaggcg 6840 aagtacgatg gtccttacag accagctact actagaccta agtcattatt gtcaagtgaa 6900 gatgttaaga gagcgtctaa taagaaaaac tcgtcttaat gcataaagaa atttattgtc 6960 aatatgacgt gtgtactcaa gggttgtgtg aatgaagtca ctgttcttgg tcacgagacg 7020 tgtagtatcg gtcatgctaa caaattgcga aagcaagttg ctgacatggt tggtgtcaca 7080 cgtaggtgtg cggaaaataa ttgtggatgg tttgtctgtg ttgttatcaa tgattttact 7140 tttgatgtgt ataattgttg tggccgtagt caccttgaaa agtgtcgtaa acgtgttgaa 7200 acaagaaatc gagaaatttg gaaacaaatt cgacgaaatc aagctgaaaa catgtctgcg 7260 acagctaaaa agtctcataa ttcgaagacc tctaagaaga aattcaaaga ggacagagaa 7320 tttgggacac caaaaagatt tttaagagat gatgttcctt tcgggattga tcgtttgttt 7380 gctttttgat tttattttat attgttatct gtttctgtgt atagactgtt tgagattggc 7440 gcttggccga ctcattgtct taccataggg gaacggactt tgtttgtgtt gttattttat 7500 ttgtatttta ttaaaattct caatgatctg aaaaggcctc gaggctaaga gattattggg 7560 gggtgagtaa gtacttttaa agtgatgatg gttacaaagg caaaaggggt aaaacccctc 7620 gcctacgtaa gcgttattac gcccggatcc cccggggagc tcgaattcgc tgaaatcacc 7680 agtctctctc tacaaatcta tctctctcta ttttttccat aaataatgtg tgagtagttt 7740 cccgataagg gaaattaggg ttcttatagg gtttcgctca tgtgttgagc atataagaaa 7800 cccttagtat gtatttgtat ttgtaaaata cttctattat caataaaatt tctaattcct 7860 aaaaccaaaa tccagtacta aaatccagat ctcctaaagt ccctatagat ctttgtcgtg 7920 aatataaacc agacacgaga cgactaaacc tggagcccag acgccgttcg aagctagaag 7980 taccgcttag gcaggaggcc gttagggaaa agatgctaag gcagggttgg ttacgttgac 8040 tcccccgtag gtttggttta aatatgatga agtggacgga aggaaggagg aagacaagga 8100 aggataaggt tgcaggccct gtgcaaggta agaagatgga aatttgatag aggtacgcta 8160 ctatacttat actatacgct aagggaatgc ttgtatttat accctatacc ccctaataac 8220 cccttatcaa tttaagaaat aatccgcata agcccccgct taaaaattgg tatcagagcc 8280 atgaataggt ctatgaccaa aactcaagag gataaaacct caccaaaata cgaaagagtt 8340 cttaactcta aagataaaag atctttcaag atcaaaacta gttccctcac accggagcat 8400 gcgatatcct cgacctgcag gcatgcaagc ttggcgtaat catggtcata gctgtttcct 8460 gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt 8520 aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc 8580 gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg 8640 agaggcggtt tgcgtattgg gccaaagaca aaagggcgac attcaaccga ttgagggagg 8700 gaaggtaaat attgacggaa attattcatt aaaggtgaat tatcaccgtc accgacttga 8760 gccatttggg aattagagcc agcaaaatca ccagtagcac cattaccatt agcaaggccg 8820 gaaacgtcac caatgaaacc atcgatagca gcaccgtaat cagtagcgac agaatcaagt 8880 ttgcctttag cgtcagactg tagcgcgttt tcatcggcat tttcggtcat agccccctta 8940 ttagcgtttg ccatcttttc ataatcaaaa tcaccggaac cagagccacc accggaaccg 9000 cctccctcag agccgccacc ctcagaaccg ccaccctcag agccaccacc ctcagagccg 9060 ccaccagaac caccaccaga gccgccgcca gcattgacag gaggcccgat ctagtaacat 9120 agatgacacc gcgcgcgata atttatccta gtttgcgcgc tatattttgt tttctatcgc 9180 gtattaaatg tataattgcg ggactctaat cataaaaacc catctcataa ataacgtcat 9240 gcattacatg ttaattatta catgcttaac gtaattcaac agaaattata tgataatcat 9300 cgcaagaccg gcaacaggat tcaatcttaa gaaactttat tgccaaatgt ttgaacgatc 9360 ggggatcatc cgggtctgtg gcgggaactc cacgaaaata tccgaacgca gcaagatatc 9420 gcggtgcatc tcggtcttgc ctgggcagtc gccgccgacg ccgttgatgt ggacgccggg 9480 cccgatcata ttgtcgctca ggatcgtggc gttgtgcttg tcggccgttg ctgtcgtaat 9540 gatatcggca ccttcgaccg cctgttccgc agagatcccg tgggcgaaga actccagcat 9600 gagatccccg cgctggagga tcatccagcc ggcgtcccgg aaaacgattc cgaagcccaa 9660 cctttcatag aaggcggcgg tggaatcgaa atctcgtgat ggcaggttgg gcgtcgcttg 9720 gtcggtcatt tcgaacccca gagtcccgct cagaagaact cgtcaagaag gcgatagaag 9780 gcgatgcgct gcgaatcggg agcggcgata ccgtaaagca cgaggaagcg gtcagcccat 9840 tcgccgccaa gctcttcagc aatatcacgg gtagccaacg ctatgtcctg atagcggtcc 9900 gccacaccca gccggccaca gtcgatgaat ccagaaaagc ggccattttc caccatgata 9960 ttcggcaagc aggcatcgcc atgggtcacg acgagatcat cgccgtcggg catgcgcgcc 10020 ttgagcctgg cgaacagttc ggctggcgcg agcccctgat gctcttcgtc cagatcatcc 10080 tgatcgacaa gaccggcttc catccgagta cgtgctcgct cgatgcgatg tttcgcttgg 10140 tggtcgaatg ggcaggtagc cggatcaagc gtatgcagcc gccgcattgc atcagccatg 10200 atggatactt tctcggcagg agcaaggtga gatgacagga gatcctgccc cggcacttcg 10260 cccaatagca gccagtccct tcccgcttca gtgacaacgt cgagcacagc tgcgcaagga 10320 acgcccgtcg tggccagcca cgatagccgc gctgcctcgt cctgcagttc attcagggca 10380 ccggacaggt cggtcttgac aaaaagaacc gggcgcccct gcgctgacag ccggaacacg 10440 gcggcatcag agcagccgat tgtctgttgt gcccagtcat agccgaatag cctctccacc 10500 caagcggccg gagaacctgc gtgcaatcca tcttgttcaa tcatgcgaaa cgatccagat 10560 ccggtgcaga ttatttggat tgagagtgaa tatgagactc taattggata ccgaggggaa 10620 tttatggaac gtcagtggag catttttgac aagaaatatt tgctagctga tagtgacctt 10680 aggcgacttt tgaacgcgca ataatggttt ctgacgtatg tgcttagctc attaaactcc 10740 agaaacccgc ggctgagtgg ctccttcaac gttgcggttc tgtcagttcc aaacgtaaaa 10800 cggcttgtcc cgcgtcatcg gcgggggtca taacgtgact cccttaattc tccgctcatg 10860 atcagattgt cgtttcccgc cttcagttta aactatcagt gtttgacagg atatattggc 10920 gggtaaacct aagagaaaag agcgtttatt agaataatcg gatatttaaa agggcgtgaa 10980 aaggtttatc cgttcgtcca tttgtatgtg catgccaacc acagggttcc ccagatctgg 11040 cgccggccag cgagacgagc aagattggcc gccgcccgaa acgatccgac agcgcgccca 11100 gcacaggtgc gcaggcaaat tgcaccaacg catacagcgc cagcagaatg ccatagtggg 11160 cggtgacgtc gttcgagtga accagatcgc gcaggaggcc cggcagcacc ggcataatca 11220 ggccgatgcc gacagcgtcg agcgcgacag tgctcagaat tacgatcagg ggtatgttgg 11280 gtttcacgtc tggcctccgg accagcctcc gctggtccga ttgaacgcgc ggattcttta 11340 tcactgataa gttggtggac atattatgtt tatcagtgat aaagtgtcaa gcatgacaaa 11400 gttgcagccg aatacagtga tccgtgccgc cctggacctg ttgaacgagg tcggcgtaga 11460 cggtctgacg acacgcaaac tggcggaacg gttgggggtt cagcagccgg cgctttactg 11520 gcacttcagg aacaagcggg cgctgctcga cgcactggcc gaagccatgc tggcggagaa 11580 tcatacgcat tcggtgccga gagccgacga cgactggcgc tcatttctga tcgggaatgc 11640 ccgcagcttc aggcaggcgc tgctcgccta ccgcgatggc gcgcgcatcc atgccggcac 11700 gcgaccgggc gcaccgcaga tggaaacggc cgacgcgcag cttcgcttcc tctgcgaggc 11760 gggtttttcg gccggggacg ccgtcaatgc gctgatgaca atcagctact tcactgttgg 11820 ggccgtgctt gaggagcagg ccggcgacag cgatgccggc gagcgcggcg gcaccgttga 11880 acaggctccg ctctcgccgc tgttgcgggc cgcgatagac gccttcgacg aagccggtcc 11940 ggacgcagcg ttcgagcagg gactcgcggt gattgtcgat ggattggcga aaaggaggct 12000 cgttgtcagg aacgttgaag gaccgagaaa gggtgacgat tgatcaggac cgctgccgga 12060 gcgcaaccca ctcactacag cagagccatg tagacaacat cccctccccc tttccaccgc 12120 gtcagacgcc cgtagcagcc cgctacgggc tttttcatgc cctgccctag cgtccaagcc 12180 tcacggccgc gctcggcctc tctggcggcc ttctggcgct cttccgcttc ctcgctcact 12240 gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta 12300 atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag 12360 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc 12420 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta 12480 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg 12540 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct tttccgctgc 12600 ataaccctgc ttcggggtca ttatagcgat tttttcggta tatccatcct ttttcgcacg 12660 atatacagga ttttgccaaa gggttcgtgt agactttcct tggtgtatcc aacggcgtca 12720 gccgggcagg ataggtgaag taggcccacc cgcgagcggg tgttccttct tcactgtccc 12780 ttattcgcac ctggcggtgc tcaacgggaa tcctgctctg cgaggctggc cggctaccgc 12840 cggcgtaaca gatgagggca agcggatggc tgatgaaacc aagccaacca ggaagggcag 12900 cccacctatc aaggtgtact gccttccaga cgaacgaaga gcgattgagg aaaaggcggc 12960 ggcggccggc atgagcctgt cggcctacct gctggccgtc ggccagggct acaaaatcac 13020 gggcgtcgtg gactatgagc acgtccgcga gctggcccgc atcaatggcg acctgggccg 13080 cctgggcggc ctgctgaaac tctggctcac cgacgacccg cgcacggcgc ggttcggtga 13140 tgccacgatc ctcgccctgc tggcgaagat cgaagagaag caggacgagc ttggcaaggt 13200 catgatgggc gtggtccgcc cgagggcaga gccatgactt ttttagccgc taaaacggcc 13260 ggggggtgcg cgtgattgcc aagcacgtcc ccatgcgctc catcaagaag agcgacttcg 13320 cggagctggt gaagtacatc accgacgagc aaggcaagac cgagcgcctt tgcgacgctc 13380 accgggctgg ttgccctcgc cgctgggctg gcggccgtct atggccctgc aaacgcgcca 13440 gaaacgccgt cgaagccgtg tgcgagacac cgcggccgcc ggcgttgtgg atacctcgcg 13500 gaaaacttgg ccctcactga cagatgaggg gcggacgttg acacttgagg ggccgactca 13560 cccggcgcgg cgttgacaga tgaggggcag gctcgatttc ggccggcgac gtggagctgg 13620 ccagcctcgc aaatcggcga aaacgcctga ttttacgcga gtttcccaca gatgatgtgg 13680 acaagcctgg ggataagtgc cctgcggtat tgacacttga ggggcgcgac tactgacaga 13740 tgaggggcgc gatccttgac acttgagggg cagagtgctg acagatgagg ggcgcaccta 13800 ttgacatttg aggggctgtc cacaggcaga aaatccagca tttgcaaggg tttccgcccg 13860 tttttcggcc accgctaacc tgtcttttaa cctgctttta aaccaatatt tataaacctt 13920 gtttttaacc agggctgcgc cctgtgcgcg tgaccgcgca cgccgaaggg gggtgccccc 13980 ccttctcgaa ccctcccggc ccgctaacgc gggcctccca tccccccagg ggctgcgccc 14040 ctcggccgcg aacggcctca ccccaaaaat ggcagcgctg gcagtccttg ccattgccgg 14100 gatcggggca gtaacgggat gggcgatcag cccgagcgcg acgcccggaa gcattgacgt 14160 gccgcaggtg ctggcatcga cattcagcga ccaggtgccg ggcagtgagg gcggcggcct 14220 gggtggcggc ctgcccttca cttcggccgt cggggcattc acggacttca tggcggggcc 14280 ggcaattttt accttgggca ttcttggcat agtggtcgcg ggtgccgtgc tcgtgttcgg 14340 gggtgcgata aacccagcga accatttgag gtgataggta agattatacc gaggtatgaa 14400 aacgagaatt ggacctttac agaattactc tatgaagcgc catatttaaa aagctaccaa 14460 gacgaagagg atgaagagga tgaggaggca gattgccttg aatatattga caatactgat 14520 aagataatat atcttttata tagaagatat cgccgtatgt aaggatttca gggggcaagg 14580 cataggcagc gcgcttatca atatatctat agaatgggca aagcataaaa acttgcatgg 14640 actaatgctt gaaacccagg acaataacct tatagcttgt aaattctatc ataattgggt 14700 aatgactcca acttattgat agtgttttat gttcagataa tgcccgatga ctttgtcatg 14760 cagctccacc gattttgaga acgacagcga cttccgtccc agccgtgcca ggtgctgcct 14820 cagattcagg ttatgccgct caattcgctg cgtatatcgc ttgctgatta cgtgcagctt 14880 tcccttcagg cgggattcat acagcggcca gccatccgtc atccatatca ccacgtcaaa 14940 gggtgacagc aggctcataa gacgccccag cgtcgccata gtgcgttcac cgaatacgtg 15000 cgcaacaacc gtcttccgga gactgtcata cgcgtaaaac agccagcgct ggcgcgattt 15060 agccccgaca tagccccact gttcgtccat ttccgcgcag acgatgacgt cactgcccgg 15120 ctgtatgcgc gaggttaccg actgcggcct gagtttttta agtgacgtaa aatcgtgttg 15180 aggccaacgc ccataatgcg ggctgttgcc cggcatccaa cgccattcat ggccatatca 15240 atgattttct ggtgcgtacc gggttgagaa gcggtgtaag tgaactgcag ttgccatgtt 15300 ttacggcagt gagagcagag atagcgctga tgtccggcgg tgcttttgcc gttacgcacc 15360 accccgtcag tagctgaaca ggagggacag ctgatagaca cagaagccac tggagcacct 15420 caaaaacacc atcatacact aaatcagtaa gttggcagca tcacccataa ttgtggtttc 15480 aaaatcggct ccgtcgatac tatgttatac gccaactttg aaaacaactt tgaaaaagct 15540 gttttctggt atttaaggtt ttagaatgca aggaacagtg aattggagtt cgtcttgtta 15600 taattagctt cttggggtat ctttaaatac tgtagaaaag aggaaggaaa taataaatgg 15660 ctaaaatgag aatatcaccg gaattgaaaa aactgatcga aaaataccgc tgcgtaaaag 15720 atacggaagg aatgtctcct gctaaggtat ataagctggt gggagaaaat gaaaacctat 15780 atttaaaaat gacggacagc cggtataaag ggaccaccta tgatgtggaa cgggaaaagg 15840 acatgatgct atggctggaa ggaaagctgc ctgttccaaa ggtcctgcac tttgaacggc 15900 atgatggctg gagcaatctg ctcatgagtg aggccgatgg cgtcctttgc tcggaagagt 15960 atgaagatga acaaagccct gaaaagatta tcgagctgta tgcggagtgc atcaggctct 16020 ttcactccat cgacatatcg gattgtccct atacgaatag cttagacagc cgcttagccg 16080 aattggatta cttactgaat aacgatctgg ccgatgtgga ttgcgaaaac tgggaagaag 16140 acactccatt taaagatccg cgcgagctgt atgatttttt aaagacggaa aagcccgaag 16200 aggaacttgt cttttcccac ggcgacctgg gagacagcaa catctttgtg aaagatggca 16260 aagtaagtgg ctttattgat cttgggagaa gcggcagggc ggacaagtgg tatgacattg 16320 ccttctgcgt ccggtcgatc agggaggata tcggggaaga acagtatgtc gagctatttt 16380 ttgacttact ggggatcaag cctgattggg agaaaataaa atattatatt ttactggatg 16440 aattgtttta gtacctagat gtggcgcaac gatgccggcg acaagcagga gcgcaccgac 16500 ttcttccgca tcaagtgttt tggctctcag gccgaggccc acggcaagta tttgggcaag 16560 gggtcgctgg tattcgtgca gggcaagatt cggaatacca agtacgagaa ggacggccag 16620 acggtctacg ggaccgactt cattgccgat aaggtggatt atctggacac caaggcacca 16680 ggcgggtcaa atcaggaata agggcacatt gccccggcgt gagtcggggc aatcccgcaa 16740 ggagggtgaa tgaatcggac gtttgaccgg aaggcataca ggcaagaact gatcgacgcg 16800 gggttttccg ccgaggatgc cgaaaccatc gcaagccgca ccgtcatgcg tgcgccccgc 16860 gaaaccttcc agtccgtcgg ctcgatggtc cagcaagcta cggccaagat cgagcgcgac 16920 agcgtgcaac tggctccccc tgccctgccc gcgccatcgg ccgccgtgga gcgttcgcgt 16980 cgtctcgaac aggaggcggc aggtttggcg aagtcgatga ccatcgacac gcgaggaact 17040 atgacgacca agaagcgaaa aaccgccggc gaggacctgg caaaacaggt cagcgaggcc 17100 aagcaggccg cgttgctgaa acacacgaag cagcagatca aggaaatgca gctttccttg 17160 ttcgatattg cgccgtggcc ggacacgatg cgagcgatgc caaacgacac ggcccgctct 17220 gccctgttca ccacgcgcaa caagaaaatc ccgcgcgagg cgctgcaaaa caaggtcatt 17280 ttccacgtca acaaggacgt gaagatcacc tacaccggcg tcgagctgcg ggccgacgat 17340 gacgaactgg tgtggcagca ggtgttggag tacgcgaagc gcacccctat cggcgagccg 17400 atcaccttca cgttctacga gctttgccag gacctgggct ggtcgatcaa tggccggtat 17460 tacacgaagg ccgaggaatg cctgtcgcgc ctacaggcga cggcgatggg cttcacgtcc 17520 gaccgcgttg ggcacctgga atcggtgtcg ctgctgcacc gcttccgcgt cctggaccgt 17580 ggcaagaaaa cgtcccgttg ccaggtcctg atcgacgagg aaatcgtcgt gctgtttgct 17640 ggcgaccact acacgaaatt catatgggag aagtaccgca agctgtcgcc gacggcccga 17700 cggatgttcg actatttcag ctcgcaccgg gagccgtacc cgctcaagct ggaaaccttc 17760 cgcctcatgt gcggatcgga ttccacccgc gtgaagaagt ggcgcgagca ggtcggcgaa 17820 gcctgcgaag agttgcgagg cagcggcctg gtggaacacg cctgggtcaa tgatgacctg 17880 gtgcattgca aacgctaggg ccttgtgggg tcagttccgg ctgggggttc agcagccagc 17940 gctttactgg catttcagga acaagcgggc actgctcgac gcacttgctt cgctcagtat 18000 cgctcgggac gcacggcgcg ctctacgaac tgccgataaa cagaggatta aaattgacaa 18060 ttgtgattaa ggctcagatt cgacggcttg gagcggccga cgtgcaggat ttccgcgaga 18120 tccgattgtc ggccctgaag aaagctccag agatgttcgg gtccgtttac gagcacgagg 18180 agaaaaagcc catggaggcg ttcgctgaac ggttgcgaga tgccgtggca ttcggcgcct 18240 acatcgacgg cgagatcatt gggctgtcgg tcttcaaaca ggaggacggc cccaaggacg 18300 ctcacaaggc gcatctgtcc ggcgttttcg tggagcccga acagcgaggc cgaggggtcg 18360 ccggtatgct gctgcgggcg ttgccggcgg gtttattgct cgtgatgatc gtccgacaga 18420 ttccaacggg aatctggtgg atgcgcatct tcatcctcgg cgcacttaat atttcgctat 18480 tctggagctt gttgtttatt tcggtctacc gcctgccggg cggggtcgcg gcgacggtag 18540 gcgctgtgca gccgctgatg gtcgtgttca tctctgccgc tctgctaggt agcccgatac 18600 gattgatggc ggtcctgggg gctatttgcg gaactgcggg cgtggcgctg ttggtgttga 18660 caccaaacgc agcgctagat cctgtcggcg tcgcagcggg cctggcgggg gcggtttcca 18720 tggcgttcgg aaccgtgctg acccgcaagt ggcaacctcc cgtgcctctg ctcaccttta 18780 ccgcctggca actggcggcc ggaggacttc tgctcgttcc agtagcttta gtgtttgatc 18840 cgccaatccc gatgcctaca ggaaccaatg ttctcggcct ggcgtggctc ggcctgatcg 18900 gagcgggttt aacctacttc ctttggttcc gggggatctc gcgactcgaa cctacagttg 18960 tttccttact gggctttctc agccccagat ctggggtcga tcagccgggg atgcatcagg 19020 ccgacagtcg gaacttcggg tccccgacct gtaccattcg gtgagcaatg gataggggag 19080 ttgatatcgt caacgttcac ttctaaagaa atagcgccac tcagcttcct cagcggcttt 19140 atccagcgat ttcctattat gtcggcatag ttctcaagat cgacagcctg tcacggttaa 19200 gcgagaaatg aataagaagg ctgataattc ggatctctgc gagggagatg atatttgatc 19260 acaggcagca acgctctgtc atcgttacaa tcaacatgct accctccgcg agatcatccg 19320 tgtttcaaac ccggcagctt agttgccgtt cttccgaata gcatcggtaa catgagcaaa 19380 gtctgccgcc ttacaacggc tctcccgctg acgccgtccc ggactgatgg gctgcctgta 19440 tcgagtggtg attttgtgcc gagctgccgg tcggggagct gttggctggc tggtggcagg 19500 atatattgtg gtgtaaacaa attgacgctt agacaactta ataacacatt gcggacgttt 19560 ttaatgtact ggggtggttt ttcttttcac cagtgagacg ggcaacagct gattgccctt 19620 caccgcctgg ccctgagaga gttgcagcaa gcggtccacg ctggtttgcc ccagcaggcg 19680 aaaatcctgt ttgatggtgg ttccgaaatc ggcaaaatcc cttataaatc aaaagaatag 19740 cccgagatag ggttgagtgt tgttccagtt tggaacaaga gtccactatt aaagaacgtg 19800 gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg atggcccact acgtgaacca 19860 tcacccaaat caagtttttt ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa 19920 gggagccccc gatttagagc ttgacgggga aagccggcga acgtggcgag aaaggaaggg 19980 aagaaagcga aaggagcggg cgccattcag gctgcgcaac tgttgggaag ggcgatcggt 20040 gcgggcctct tcgctattac gccagctggc gaaaggggga tgtgctgcaa ggcgattaag 20100 ttgggtaacg ccagggtttt cccagtcacg acgttgtaaa acgacggcca gtgaattcga 20160 gctcggtacc cccc 20174 2 19705 DNA Artificial Sequence Description of Artificial Sequence pBTA-delta -MP 2 tactccaaaa atgtcaaaga tacagtctca gaagaccaaa gggctattga gacttttcaa 60 caaagggtaa tttcgggaaa cctcctcgga ttccattgcc cagctatctg tcacttcatc 120 gaaaggacag tagaaaagga aggtggctcc tacaaatgcc atcattgcga taaaggaaag 180 gctatcattc aagatgcctc tgccgacagt ggtcccaaag atggaccccc acccacgagg 240 agcatcgtgg aaaaagaaga cgtcccaacc acgtcttcaa agcaagtgga ttgatgtgac 300 atctccactg acgtaaggga tgacgcacaa tcccactatc cttcgcaaga cccttcttct 360 atataaggaa gttcatttca tttggagagg acagcccaag ctttctagag gatccataaa 420 acatttcaat cctttgaacg cggtagaacg tgctaattgg attttggtga gaacgcggta 480 gaacgtactt atcacctaca gttttatttt gtttttcttt ttggtttaat ctatccagct 540 tagtaccgag tgggggaaag tgactggtgt gcctaaaacc ttttctttga tactttgtaa 600 aaatacatac agatacaatg gcgaacggta acttcaagtt gtctcaattg ctcaatgtgg 660 acgagatgtc tgctgagcag aggagtcatt tctttgactt gatgctgact aaacctgatt 720 gtgagatcgg gcaaatgatg caaagagttg ttgttgataa agtcgatgac atgattagag 780 aaagaaagac taaagatcca gtgattgttc atgaagttct ttctcagaag gaacagaaca 840 agttgatgga aatttatcct gaattcaata tcgtgtttaa agacgacaaa aacatggttc 900 atgggtttgc ggctgctgag cgaaaactac aagctttatt gcttttagat agagttcctg 960 ctctgcaaga ggtggatgac atcggtggtc aatggtcgtt ttgggtaact agaggtgaga 1020 aaaggattca ttcctgttgt ccaaatctag atattcggga tgatcagaga gaaatttctc 1080 gacagatatt tcttactgct attggtgatc aagctagaag tggtaagaga cagatgtcgg 1140 agaatgagct gtggatgtat gaccaatttc gtgaaaatat tgctgcgcct aacgcggtta 1200 ggtgcaataa tacatatcag ggttgtacat gtaggggttt ttctgatggt aagaagaaag 1260 gcgcgcagta tgcgatagct cttcacagcc tgtatgactt caagttgaaa gacttgatgg 1320 ctactatggt tgagaagaaa actaaagtgg ttcatgctgc tatgcttttt gctcctgaaa 1380 gtatgttagt ggacgaaggt ccattacctt ctgttgacgg ttactacatg aagaagaacg 1440 ggaagatcta tttcggtttt gagaaagatc cttccttttc ttacattcat gactgggaag 1500 agtacaagaa gtatctactg gggaagccag tgagttacca agggaatgtg ttctacttcg 1560 aaccgtggca ggtgagagga gacacaatgc ttttttcgat ctacaggata gctggagttc 1620 cgaggaggtc tctatcatcg caagagtact accgaagaat atatatcagt agatgggaaa 1680 gcatggttgt tgtcccaatt ttcgatctgg tcgaatcaac gcgagagttg gtcaagaaag 1740 acctgtttgt agagaaacaa ttcatggaca agtgtttgga ttacatagct aggttatctg 1800 accagcagct gaccataagc aatgttaaat catacttgag ttcaaataat tgggtcttat 1860 tcataaacgg ggcggccgtg aagaacaagc aaagtgtaga ttctcgagat ttacagttgt 1920 tggctcaaac tttgctagtg aaggaacaag tggcgagacc tgtcatgagg gagttgcgtg 1980 aagcaattct gactgagacg aaacctatca cgtcattgac tgatgtgctg ggtttaatat 2040 caagaaaact gtggaagcag tttgctaaca agatcgcagt cggcggattc gttggcatgg 2100 ttggtactct aattggattc tatccaaaga aggtactaac ctgggcgaag gacacaccaa 2160 atggtccaga actatgttac gagaactcgc acaaaaccaa ggtgatagta tttctgagtg 2220 ttgtgtatgc cattggagga atcacgctta tgcgtcgaga catccgagat ggactggtga 2280 aaaaactatg tgatatgttt gatatcaaac ggggggccca tgtcttagac gttgagaatc 2340 cgtgccgcta ttatgaaatc aacgatttct ttagcagtct gtattcggca tctgagtccg 2400 gtgagaccgt tttaccagat ttatccgagg taaaagccaa gtctgataag ctattgcagc 2460 agaagaaaga aatcgctgac gagtttctaa gtgcaaaatt ctctaactat tctggcagtt 2520 cggtgagaac ttctccacca tcggtggtcg gttcatctcg aagcggactg ggtctgttgt 2580 tggaagacag taacgtgctg acccaagcta gagttggagt ttcaagaaag gtagacgatg 2640 aggagatcat ggagcagttt ctgagtggtc ttattgacac tgaagcagaa attgacgagg 2700 ttgtttcagc cttttcagct gaatgtgaaa gaggggaaac aagcggtaca aaggtgttgt 2760 gtaaaccttt aacgccacca ggatttgaga acgtgttgcc agctgtcaaa cctttggtca 2820 gcaaaggaaa aacggtcaaa cgtgtcgatt acttccaagt gatgggaggt gagagattac 2880 caaaaaggcc ggttgtcagt ggagacgatt ctgtggacgc tagaagagag tttctgtact 2940 acttagatgc ggagagagtc gctcaaaatg atgaaattat gtctctgtat cgtgactatt 3000 cgagaggagt tattcgaact ggaggtcaga attacccgca cggactggga gtgtgggatg 3060 tggagatgaa gaactggtgc atacgtccag tggtcactga acatgcttat gtgttccaac 3120 cagacaaacg tatggatgat tggtcgggat acttagaagt ggctgtttgg gaacgaggta 3180 tgttggtcaa cgacttcgcg gtcgaaagga tgagtgatta tgtcatagtt tgcgatcaga 3240 cgtatctttg caataacagg taataatcct ctctcttgat atttttaaat tatagaatta 3300 attagtttac tttattcttt actatatgat ttaaatagtt taatcttgtt tttgagtaaa 3360 ctattcgatt ttgatatttg tattcgtcct acaaagttgg aaatactgat gatattttct 3420 tttgaacgtg atacctacca atactaatct tacggaatct tttaatagag cactaatcaa 3480 catggaacta aagaccaatt cttaagtgtc tctgttgtac agttcatttt agtagtgcgt 3540 ttaagtatta ttatctccct tcatgcgggg caattatgta gattaaaatc gaaattatat 3600 aaaatttaca taagtctaag tctagggtct ccagctaatt gttatttttt taacgatgtt 3660 gactaaagca ataacgacgt tgacttgtgt taaacaggtt gatcttggac aatttaagtg 3720 ccctggatct aggaccagtt aactgttctt ttgaattagt tgacggtgta cctggttgtg 3780 gtaagtcgac aatgattgtc aactcagcta atccttgtgt cgatgtggtt ctctctactg 3840 ggagagcagc aaccgacgac ttgatcgaga gattcgcgag caaaggtttt ccatgcaaat 3900 tgaaaaggag agtgaagacg gttgattctt ttttgatgca ttgtgtcgat ggttctttaa 3960 ccggagacgt gttgcatttc gacgaagctc tcatggccca tgctggtatg gtgtactttt 4020 gcgctcagat agctggtgct aaacgatgta tctgtcaagg agatcagaat caaatttctt 4080 tcaagcctag ggtatctcaa gttgatttga ggttttctag tctggtcgga aagtttgaca 4140 ttgttacaga aaaaagagaa acttacagaa gtccagcaga tgtggctgcc gtattgaaca 4200 agtactatac tggagatgtc agaacacata acgcgactgc taattcgatg acggtgagga 4260 agattgtgtc taaagaacag gtttctttga agcctggtgc tcagtacata actttccttc 4320 agtctgagaa gaaggagttg gtaaatttgt tggcattgag gaaagtggca gctaaagtga 4380 gtacagtaca cgagtcgcaa ggagagacat tcaaagatgt agtcctagtc aggacgaaac 4440 ctacggatga ctcaatcgct agaggtcggg agtacttaat cgtggcattg tcgcgtcaca 4500 cacaatcact tgtgtatgaa actgtgaaag aggacgatgt aagcaaagag atcagggaaa 4560 gtgccgcgct tacgaaggcg gctttggcaa gattttttgt tactgagacc gtcttatgac 4620 ggtttcggtc taggtttgat gtctttagac atcatgaagg gccttgcgcc gttccagatt 4680 caggtacgat tacggacttg gagatgtggt acgacgcttt gtttccggga aattcgttaa 4740 gagactcaag cctagacggg tatttggtgg caacgactga ttgcaatttg cgattagaca 4800 atgttacgat caaaagtgga aactggaaag acaagtttgc tgaaaaagaa acgtttctga 4860 aaccggttat tcgtactgct atgcctgaca aaaggaagac tactcagttg gagagtttgt 4920 tagcattgca gaaaaggaac caagcggcac ccgatctaca agaaaatgtg cacgcgacag 4980 ttctaatcga agagacgatg aagaagctga aatctgttgt ctacgatgtg ggaaaaattc 5040 gggctgatcc tattgtcaat agagctcaaa tggagagatg gtggagaaat caaagcacag 5100 cggtacaggc taaggtagta gcagatgtga gagagttaca tgaaatagac tattcgtctt 5160 acatgtatat gatcaaatct gacgtgaaac ctaagactga tttaacaccg caatttgaat 5220 actcagctct acagactgtt gtgtatcacg agaagttgat caactcgttg ttcggtccaa 5280 ttttcaaaga aattaatgaa cgcaagttgg atgctatgca accacatttt gtgttcaaca 5340 cgagaatgac atcgagtgat ttaaacgatc gagtgaagtt cttaaatacg gaagcggctt 5400 acgactttgt tgagatagac atgtctaaat tcgacaagtc ggcaaatcgc ttccatttac 5460 aactgcagct ggagatttac aggttatttg ggctggatga gtgggcggcc ttcctttggg 5520 aggtgtcgca cactcaaact actgtgagag atattcaaaa tggtatgatg gcgcatattt 5580 ggtaccaaca aaagagtgga gatgctgata cttataatgc aaattcagat agaacactgt 5640 gtgcactctt gtctgaatta ccattggaga aagcagtcat ggttacatat ggaggagatg 5700 actcactgat tgcgtttcct agaggaacgc agtttgttga tccgtgtcca aagttggcta 5760 ctaagtggaa tttcgagtgc aagattttta agtacgatgt cccaatgttt tgtgggaagt 5820 tcttgcttaa gacgtcatcg tgttacgagt tcgtgccaga tccggtaaaa gttctgacga 5880 agttggggaa aaagagtata aaggatgtgc aacatttagc cgagatctac atctcgctga 5940 atgattccaa tagagctctt gggaactaca tggtggtatc caaactgtcc gagtctgttt 6000 cagaccggta tttgtacaaa ggtgattctg ttcatgcgct ttgtgcgcta tggaagcata 6060 ttaagagttt tacagctctg tgtacattat tccgagacga aaacgataag gaattgaacc 6120 cggctaaggt tgattggaag aaggcacaga gagctgtgtc aaacttttac gactggtaat 6180 atggaagaca agtcattggt caccttgaag aagaagactg gcgcgccacg tgttaattaa 6240 ctgattcgac taggcgcctc aatgtggaag aactgaacag ttcggattac attgaaggcg 6300 attttaccga tcaagaggtt ttcggtgagt tcatgtcttt gaaacaagtg gagatgaaga 6360 cgattgaggc gaagtacgat ggtccttaca gaccagctac tactagacct aagtcattat 6420 tgtcaagtga agatgttaag agagcgtcta ataagaaaaa ctcgtcttaa tgcataaaga 6480 aatttattgt caatatgacg tgtgtactca agggttgtgt gaatgaagtc actgttcttg 6540 gtcacgagac gtgtagtatc ggtcatgcta acaaattgcg aaagcaagtt gctgacatgg 6600 ttggtgtcac acgtaggtgt gcggaaaata attgtggatg gtttgtctgt gttgttatca 6660 atgattttac ttttgatgtg tataattgtt gtggccgtag tcaccttgaa aagtgtcgta 6720 aacgtgttga aacaagaaat cgagaaattt ggaaacaaat tcgacgaaat caagctgaaa 6780 acatgtctgc gacagctaaa aagtctcata attcgaagac ctctaagaag aaattcaaag 6840 aggacagaga atttgggaca ccaaaaagat ttttaagaga tgatgttcct ttcgggattg 6900 atcgtttgtt tgctttttga ttttatttta tattgttatc tgtttctgtg tatagactgt 6960 ttgagattgg cgcttggccg actcattgtc ttaccatagg ggaacggact ttgtttgtgt 7020 tgttatttta tttgtatttt attaaaattc tcaatgatct gaaaaggcct cgaggctaag 7080 agattattgg ggggtgagta agtactttta aagtgatgat ggttacaaag gcaaaagggg 7140 taaaacccct cgcctacgta agcgttatta cgcccggatc ccccggggag ctcgaattcg 7200 ctgaaatcac cagtctctct ctacaaatct atctctctct attttttcca taaataatgt 7260 gtgagtagtt tcccgataag ggaaattagg gttcttatag ggtttcgctc atgtgttgag 7320 catataagaa acccttagta tgtatttgta tttgtaaaat acttctatta tcaataaaat 7380 ttctaattcc taaaaccaaa atccagtact aaaatccaga tctcctaaag tccctataga 7440 tctttgtcgt gaatataaac cagacacgag acgactaaac ctggagccca gacgccgttc 7500 gaagctagaa gtaccgctta ggcaggaggc cgttagggaa aagatgctaa ggcagggttg 7560 gttacgttga ctcccccgta ggtttggttt aaatatgatg aagtggacgg aaggaaggag 7620 gaagacaagg aaggataagg ttgcaggccc tgtgcaaggt aagaagatgg aaatttgata 7680 gaggtacgct actatactta tactatacgc taagggaatg cttgtattta taccctatac 7740 cccctaataa ccccttatca atttaagaaa taatccgcat aagcccccgc ttaaaaattg 7800 gtatcagagc catgaatagg tctatgacca aaactcaaga ggataaaacc tcaccaaaat 7860 acgaaagagt tcttaactct aaagataaaa gatctttcaa gatcaaaact agttccctca 7920 caccggagca tgcgatatcc tcgacctgca ggcatgcaag cttggcgtaa tcatggtcat 7980 agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa 8040 gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc 8100 gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc 8160 aacgcgcggg gagaggcggt ttgcgtattg ggccaaagac aaaagggcga cattcaaccg 8220 attgagggag ggaaggtaaa tattgacgga aattattcat taaaggtgaa ttatcaccgt 8280 caccgacttg agccatttgg gaattagagc cagcaaaatc accagtagca ccattaccat 8340 tagcaaggcc ggaaacgtca ccaatgaaac catcgatagc agcaccgtaa tcagtagcga 8400 cagaatcaag tttgccttta gcgtcagact gtagcgcgtt ttcatcggca ttttcggtca 8460 tagccccctt attagcgttt gccatctttt cataatcaaa atcaccggaa ccagagccac 8520 caccggaacc gcctccctca gagccgccac cctcagaacc gccaccctca gagccaccac 8580 cctcagagcc gccaccagaa ccaccaccag agccgccgcc agcattgaca ggaggcccga 8640 tctagtaaca tagatgacac cgcgcgcgat aatttatcct agtttgcgcg ctatattttg 8700 ttttctatcg cgtattaaat gtataattgc gggactctaa tcataaaaac ccatctcata 8760 aataacgtca tgcattacat gttaattatt acatgcttaa cgtaattcaa cagaaattat 8820 atgataatca tcgcaagacc ggcaacagga ttcaatctta agaaacttta ttgccaaatg 8880 tttgaacgat cggggatcat ccgggtctgt ggcgggaact ccacgaaaat atccgaacgc 8940 agcaagatat cgcggtgcat ctcggtcttg cctgggcagt cgccgccgac gccgttgatg 9000 tggacgccgg gcccgatcat attgtcgctc aggatcgtgg cgttgtgctt gtcggccgtt 9060 gctgtcgtaa tgatatcggc accttcgacc gcctgttccg cagagatccc gtgggcgaag 9120 aactccagca tgagatcccc gcgctggagg atcatccagc cggcgtcccg gaaaacgatt 9180 ccgaagccca acctttcata gaaggcggcg gtggaatcga aatctcgtga tggcaggttg 9240 ggcgtcgctt ggtcggtcat ttcgaacccc agagtcccgc tcagaagaac tcgtcaagaa 9300 ggcgatagaa ggcgatgcgc tgcgaatcgg gagcggcgat accgtaaagc acgaggaagc 9360 ggtcagccca ttcgccgcca agctcttcag caatatcacg ggtagccaac gctatgtcct 9420 gatagcggtc cgccacaccc agccggccac agtcgatgaa tccagaaaag cggccatttt 9480 ccaccatgat attcggcaag caggcatcgc catgggtcac gacgagatca tcgccgtcgg 9540 gcatgcgcgc cttgagcctg gcgaacagtt cggctggcgc gagcccctga tgctcttcgt 9600 ccagatcatc ctgatcgaca agaccggctt ccatccgagt acgtgctcgc tcgatgcgat 9660 gtttcgcttg gtggtcgaat gggcaggtag ccggatcaag cgtatgcagc cgccgcattg 9720 catcagccat gatggatact ttctcggcag gagcaaggtg agatgacagg agatcctgcc 9780 ccggcacttc gcccaatagc agccagtccc ttcccgcttc agtgacaacg tcgagcacag 9840 ctgcgcaagg aacgcccgtc gtggccagcc acgatagccg cgctgcctcg tcctgcagtt 9900 cattcagggc accggacagg tcggtcttga caaaaagaac cgggcgcccc tgcgctgaca 9960 gccggaacac ggcggcatca gagcagccga ttgtctgttg tgcccagtca tagccgaata 10020 gcctctccac ccaagcggcc ggagaacctg cgtgcaatcc atcttgttca atcatgcgaa 10080 acgatccaga tccggtgcag attatttgga ttgagagtga atatgagact ctaattggat 10140 accgagggga atttatggaa cgtcagtgga gcatttttga caagaaatat ttgctagctg 10200 atagtgacct taggcgactt ttgaacgcgc aataatggtt tctgacgtat gtgcttagct 10260 cattaaactc cagaaacccg cggctgagtg gctccttcaa cgttgcggtt ctgtcagttc 10320 caaacgtaaa acggcttgtc ccgcgtcatc ggcgggggtc ataacgtgac tcccttaatt 10380 ctccgctcat gatcagattg tcgtttcccg ccttcagttt aaactatcag tgtttgacag 10440 gatatattgg cgggtaaacc taagagaaaa gagcgtttat tagaataatc ggatatttaa 10500 aagggcgtga aaaggtttat ccgttcgtcc atttgtatgt gcatgccaac cacagggttc 10560 cccagatctg gcgccggcca gcgagacgag caagattggc cgccgcccga aacgatccga 10620 cagcgcgccc agcacaggtg cgcaggcaaa ttgcaccaac gcatacagcg ccagcagaat 10680 gccatagtgg gcggtgacgt cgttcgagtg aaccagatcg cgcaggaggc ccggcagcac 10740 cggcataatc aggccgatgc cgacagcgtc gagcgcgaca gtgctcagaa ttacgatcag 10800 gggtatgttg ggtttcacgt ctggcctccg gaccagcctc cgctggtccg attgaacgcg 10860 cggattcttt atcactgata agttggtgga catattatgt ttatcagtga taaagtgtca 10920 agcatgacaa agttgcagcc gaatacagtg atccgtgccg ccctggacct gttgaacgag 10980 gtcggcgtag acggtctgac gacacgcaaa ctggcggaac ggttgggggt tcagcagccg 11040 gcgctttact ggcacttcag gaacaagcgg gcgctgctcg acgcactggc cgaagccatg 11100 ctggcggaga atcatacgca ttcggtgccg agagccgacg acgactggcg ctcatttctg 11160 atcgggaatg cccgcagctt caggcaggcg ctgctcgcct accgcgatgg cgcgcgcatc 11220 catgccggca cgcgaccggg cgcaccgcag atggaaacgg ccgacgcgca gcttcgcttc 11280 ctctgcgagg cgggtttttc ggccggggac gccgtcaatg cgctgatgac aatcagctac 11340 ttcactgttg gggccgtgct tgaggagcag gccggcgaca gcgatgccgg cgagcgcggc 11400 ggcaccgttg aacaggctcc gctctcgccg ctgttgcggg ccgcgataga cgccttcgac 11460 gaagccggtc cggacgcagc gttcgagcag ggactcgcgg tgattgtcga tggattggcg 11520 aaaaggaggc tcgttgtcag gaacgttgaa ggaccgagaa agggtgacga ttgatcagga 11580 ccgctgccgg agcgcaaccc actcactaca gcagagccat gtagacaaca tcccctcccc 11640 ctttccaccg cgtcagacgc ccgtagcagc ccgctacggg ctttttcatg ccctgcccta 11700 gcgtccaagc ctcacggccg cgctcggcct ctctggcggc cttctggcgc tcttccgctt 11760 cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact 11820 caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag 11880 caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata 11940 ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc 12000 cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg 12060 ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc 12120 ttttccgctg cataaccctg cttcggggtc attatagcga ttttttcggt atatccatcc 12180 tttttcgcac gatatacagg attttgccaa agggttcgtg tagactttcc ttggtgtatc 12240 caacggcgtc agccgggcag gataggtgaa gtaggcccac ccgcgagcgg gtgttccttc 12300 ttcactgtcc cttattcgca cctggcggtg ctcaacggga atcctgctct gcgaggctgg 12360 ccggctaccg ccggcgtaac agatgagggc aagcggatgg ctgatgaaac caagccaacc 12420 aggaagggca gcccacctat caaggtgtac tgccttccag acgaacgaag agcgattgag 12480 gaaaaggcgg cggcggccgg catgagcctg tcggcctacc tgctggccgt cggccagggc 12540 tacaaaatca cgggcgtcgt ggactatgag cacgtccgcg agctggcccg catcaatggc 12600 gacctgggcc gcctgggcgg cctgctgaaa ctctggctca ccgacgaccc gcgcacggcg 12660 cggttcggtg atgccacgat cctcgccctg ctggcgaaga tcgaagagaa gcaggacgag 12720 cttggcaagg tcatgatggg cgtggtccgc ccgagggcag agccatgact tttttagccg 12780 ctaaaacggc cggggggtgc gcgtgattgc caagcacgtc cccatgcgct ccatcaagaa 12840 gagcgacttc gcggagctgg tgaagtacat caccgacgag caaggcaaga ccgagcgcct 12900 ttgcgacgct caccgggctg gttgccctcg ccgctgggct ggcggccgtc tatggccctg 12960 caaacgcgcc agaaacgccg tcgaagccgt gtgcgagaca ccgcggccgc cggcgttgtg 13020 gatacctcgc ggaaaacttg gccctcactg acagatgagg ggcggacgtt gacacttgag 13080 gggccgactc acccggcgcg gcgttgacag atgaggggca ggctcgattt cggccggcga 13140 cgtggagctg gccagcctcg caaatcggcg aaaacgcctg attttacgcg agtttcccac 13200 agatgatgtg gacaagcctg gggataagtg ccctgcggta ttgacacttg aggggcgcga 13260 ctactgacag atgaggggcg cgatccttga cacttgaggg gcagagtgct gacagatgag 13320 gggcgcacct attgacattt gaggggctgt ccacaggcag aaaatccagc atttgcaagg 13380 gtttccgccc gtttttcggc caccgctaac ctgtctttta acctgctttt aaaccaatat 13440 ttataaacct tgtttttaac cagggctgcg ccctgtgcgc gtgaccgcgc acgccgaagg 13500 ggggtgcccc cccttctcga accctcccgg cccgctaacg cgggcctccc atccccccag 13560 gggctgcgcc cctcggccgc gaacggcctc accccaaaaa tggcagcgct ggcagtcctt 13620 gccattgccg ggatcggggc agtaacggga tgggcgatca gcccgagcgc gacgcccgga 13680 agcattgacg tgccgcaggt gctggcatcg acattcagcg accaggtgcc gggcagtgag 13740 ggcggcggcc tgggtggcgg cctgcccttc acttcggccg tcggggcatt cacggacttc 13800 atggcggggc cggcaatttt taccttgggc attcttggca tagtggtcgc gggtgccgtg 13860 ctcgtgttcg ggggtgcgat aaacccagcg aaccatttga ggtgataggt aagattatac 13920 cgaggtatga aaacgagaat tggaccttta cagaattact ctatgaagcg ccatatttaa 13980 aaagctacca agacgaagag gatgaagagg atgaggaggc agattgcctt gaatatattg 14040 acaatactga taagataata tatcttttat atagaagata tcgccgtatg taaggatttc 14100 agggggcaag gcataggcag cgcgcttatc aatatatcta tagaatgggc aaagcataaa 14160 aacttgcatg gactaatgct tgaaacccag gacaataacc ttatagcttg taaattctat 14220 cataattggg taatgactcc aacttattga tagtgtttta tgttcagata atgcccgatg 14280 actttgtcat gcagctccac cgattttgag aacgacagcg acttccgtcc cagccgtgcc 14340 aggtgctgcc tcagattcag gttatgccgc tcaattcgct gcgtatatcg cttgctgatt 14400 acgtgcagct ttcccttcag gcgggattca tacagcggcc agccatccgt catccatatc 14460 accacgtcaa agggtgacag caggctcata agacgcccca gcgtcgccat agtgcgttca 14520 ccgaatacgt gcgcaacaac cgtcttccgg agactgtcat acgcgtaaaa cagccagcgc 14580 tggcgcgatt tagccccgac atagccccac tgttcgtcca tttccgcgca gacgatgacg 14640 tcactgcccg gctgtatgcg cgaggttacc gactgcggcc tgagtttttt aagtgacgta 14700 aaatcgtgtt gaggccaacg cccataatgc gggctgttgc ccggcatcca acgccattca 14760 tggccatatc aatgattttc tggtgcgtac cgggttgaga agcggtgtaa gtgaactgca 14820 gttgccatgt tttacggcag tgagagcaga gatagcgctg atgtccggcg gtgcttttgc 14880 cgttacgcac caccccgtca gtagctgaac aggagggaca gctgatagac acagaagcca 14940 ctggagcacc tcaaaaacac catcatacac taaatcagta agttggcagc atcacccata 15000 attgtggttt caaaatcggc tccgtcgata ctatgttata cgccaacttt gaaaacaact 15060 ttgaaaaagc tgttttctgg tatttaaggt tttagaatgc aaggaacagt gaattggagt 15120 tcgtcttgtt ataattagct tcttggggta tctttaaata ctgtagaaaa gaggaaggaa 15180 ataataaatg gctaaaatga gaatatcacc ggaattgaaa aaactgatcg aaaaataccg 15240 ctgcgtaaaa gatacggaag gaatgtctcc tgctaaggta tataagctgg tgggagaaaa 15300 tgaaaaccta tatttaaaaa tgacggacag ccggtataaa gggaccacct atgatgtgga 15360 acgggaaaag gacatgatgc tatggctgga aggaaagctg cctgttccaa aggtcctgca 15420 ctttgaacgg catgatggct ggagcaatct gctcatgagt gaggccgatg gcgtcctttg 15480 ctcggaagag tatgaagatg aacaaagccc tgaaaagatt atcgagctgt atgcggagtg 15540 catcaggctc tttcactcca tcgacatatc ggattgtccc tatacgaata gcttagacag 15600 ccgcttagcc gaattggatt acttactgaa taacgatctg gccgatgtgg attgcgaaaa 15660 ctgggaagaa gacactccat ttaaagatcc gcgcgagctg tatgattttt taaagacgga 15720 aaagcccgaa gaggaacttg tcttttccca cggcgacctg ggagacagca acatctttgt 15780 gaaagatggc aaagtaagtg gctttattga tcttgggaga agcggcaggg cggacaagtg 15840 gtatgacatt gccttctgcg tccggtcgat cagggaggat atcggggaag aacagtatgt 15900 cgagctattt tttgacttac tggggatcaa gcctgattgg gagaaaataa aatattatat 15960 tttactggat gaattgtttt agtacctaga tgtggcgcaa cgatgccggc gacaagcagg 16020 agcgcaccga cttcttccgc atcaagtgtt ttggctctca ggccgaggcc cacggcaagt 16080 atttgggcaa ggggtcgctg gtattcgtgc agggcaagat tcggaatacc aagtacgaga 16140 aggacggcca gacggtctac gggaccgact tcattgccga taaggtggat tatctggaca 16200 ccaaggcacc aggcgggtca aatcaggaat aagggcacat tgccccggcg tgagtcgggg 16260 caatcccgca aggagggtga atgaatcgga cgtttgaccg gaaggcatac aggcaagaac 16320 tgatcgacgc ggggttttcc gccgaggatg ccgaaaccat cgcaagccgc accgtcatgc 16380 gtgcgccccg cgaaaccttc cagtccgtcg gctcgatggt ccagcaagct acggccaaga 16440 tcgagcgcga cagcgtgcaa ctggctcccc ctgccctgcc cgcgccatcg gccgccgtgg 16500 agcgttcgcg tcgtctcgaa caggaggcgg caggtttggc gaagtcgatg accatcgaca 16560 cgcgaggaac tatgacgacc aagaagcgaa aaaccgccgg cgaggacctg gcaaaacagg 16620 tcagcgaggc caagcaggcc gcgttgctga aacacacgaa gcagcagatc aaggaaatgc 16680 agctttcctt gttcgatatt gcgccgtggc cggacacgat gcgagcgatg ccaaacgaca 16740 cggcccgctc tgccctgttc accacgcgca acaagaaaat cccgcgcgag gcgctgcaaa 16800 acaaggtcat tttccacgtc aacaaggacg tgaagatcac ctacaccggc gtcgagctgc 16860 gggccgacga tgacgaactg gtgtggcagc aggtgttgga gtacgcgaag cgcaccccta 16920 tcggcgagcc gatcaccttc acgttctacg agctttgcca ggacctgggc tggtcgatca 16980 atggccggta ttacacgaag gccgaggaat gcctgtcgcg cctacaggcg acggcgatgg 17040 gcttcacgtc cgaccgcgtt gggcacctgg aatcggtgtc gctgctgcac cgcttccgcg 17100 tcctggaccg tggcaagaaa acgtcccgtt gccaggtcct gatcgacgag gaaatcgtcg 17160 tgctgtttgc tggcgaccac tacacgaaat tcatatggga gaagtaccgc aagctgtcgc 17220 cgacggcccg acggatgttc gactatttca gctcgcaccg ggagccgtac ccgctcaagc 17280 tggaaacctt ccgcctcatg tgcggatcgg attccacccg cgtgaagaag tggcgcgagc 17340 aggtcggcga agcctgcgaa gagttgcgag gcagcggcct ggtggaacac gcctgggtca 17400 atgatgacct ggtgcattgc aaacgctagg gccttgtggg gtcagttccg gctgggggtt 17460 cagcagccag cgctttactg gcatttcagg aacaagcggg cactgctcga cgcacttgct 17520 tcgctcagta tcgctcggga cgcacggcgc gctctacgaa ctgccgataa acagaggatt 17580 aaaattgaca attgtgatta aggctcagat tcgacggctt ggagcggccg acgtgcagga 17640 tttccgcgag atccgattgt cggccctgaa gaaagctcca gagatgttcg ggtccgttta 17700 cgagcacgag gagaaaaagc ccatggaggc gttcgctgaa cggttgcgag atgccgtggc 17760 attcggcgcc tacatcgacg gcgagatcat tgggctgtcg gtcttcaaac aggaggacgg 17820 ccccaaggac gctcacaagg cgcatctgtc cggcgttttc gtggagcccg aacagcgagg 17880 ccgaggggtc gccggtatgc tgctgcgggc gttgccggcg ggtttattgc tcgtgatgat 17940 cgtccgacag attccaacgg gaatctggtg gatgcgcatc ttcatcctcg gcgcacttaa 18000 tatttcgcta ttctggagct tgttgtttat ttcggtctac cgcctgccgg gcggggtcgc 18060 ggcgacggta ggcgctgtgc agccgctgat ggtcgtgttc atctctgccg ctctgctagg 18120 tagcccgata cgattgatgg cggtcctggg ggctatttgc ggaactgcgg gcgtggcgct 18180 gttggtgttg acaccaaacg cagcgctaga tcctgtcggc gtcgcagcgg gcctggcggg 18240 ggcggtttcc atggcgttcg gaaccgtgct gacccgcaag tggcaacctc ccgtgcctct 18300 gctcaccttt accgcctggc aactggcggc cggaggactt ctgctcgttc cagtagcttt 18360 agtgtttgat ccgccaatcc cgatgcctac aggaaccaat gttctcggcc tggcgtggct 18420 cggcctgatc ggagcgggtt taacctactt cctttggttc cgggggatct cgcgactcga 18480 acctacagtt gtttccttac tgggctttct cagccccaga tctggggtcg atcagccggg 18540 gatgcatcag gccgacagtc ggaacttcgg gtccccgacc tgtaccattc ggtgagcaat 18600 ggatagggga gttgatatcg tcaacgttca cttctaaaga aatagcgcca ctcagcttcc 18660 tcagcggctt tatccagcga tttcctatta tgtcggcata gttctcaaga tcgacagcct 18720 gtcacggtta agcgagaaat gaataagaag gctgataatt cggatctctg cgagggagat 18780 gatatttgat cacaggcagc aacgctctgt catcgttaca atcaacatgc taccctccgc 18840 gagatcatcc gtgtttcaaa cccggcagct tagttgccgt tcttccgaat agcatcggta 18900 acatgagcaa agtctgccgc cttacaacgg ctctcccgct gacgccgtcc cggactgatg 18960 ggctgcctgt atcgagtggt gattttgtgc cgagctgccg gtcggggagc tgttggctgg 19020 ctggtggcag gatatattgt ggtgtaaaca aattgacgct tagacaactt aataacacat 19080 tgcggacgtt tttaatgtac tggggtggtt tttcttttca ccagtgagac gggcaacagc 19140 tgattgccct tcaccgcctg gccctgagag agttgcagca agcggtccac gctggtttgc 19200 cccagcaggc gaaaatcctg tttgatggtg gttccgaaat cggcaaaatc ccttataaat 19260 caaaagaata gcccgagata gggttgagtg ttgttccagt ttggaacaag agtccactat 19320 taaagaacgt ggactccaac gtcaaagggc gaaaaaccgt ctatcagggc gatggcccac 19380 tacgtgaacc atcacccaaa tcaagttttt tggggtcgag gtgccgtaaa gcactaaatc 19440 ggaaccctaa agggagcccc cgatttagag cttgacgggg aaagccggcg aacgtggcga 19500 gaaaggaagg gaagaaagcg aaaggagcgg gcgccattca ggctgcgcaa ctgttgggaa 19560 gggcgatcgg tgcgggcctc ttcgctatta cgccagctgg cgaaaggggg atgtgctgca 19620 aggcgattaa gttgggtaac gccagggttt tcccagtcac gacgttgtaa aacgacggcc 19680 agtgaattcg agctcggtac ccccc 19705 3 6411 DNA Artificial Sequence Description of Artificial Sequence pBTA-delta-MP-delta-16K 3 gatccataaa acatttcaat cctttgaacg cggtagaacg tgctaattgg attttggtga 60 gaacgcggta gaacgtactt atcacctaca gttttatttt gtttttcttt ttggtttaat 120 ctatccagct tagtaccgag tgggggaaag tgactggtgt gcctaaaacc ttttctttga 180 tactttgtaa aaatacatac agatacaatg gcgaacggta acttcaagtt gtctcaattg 240 ctcaatgtgg acgagatgtc tgctgagcag aggagtcatt tctttgactt gatgctgact 300 aaacctgatt gtgagatcgg gcaaatgatg caaagagttg ttgttgataa agtcgatgac 360 atgattagag aaagaaagac taaagatcca gtgattgttc atgaagttct ttctcagaag 420 gaacagaaca agttgatgga aatttatcct gaattcaata tcgtgtttaa agacgacaaa 480 aacatggttc atgggtttgc ggctgctgag cgaaaactac aagctttatt gcttttagat 540 agagttcctg ctctgcaaga ggtggatgac atcggtggtc aatggtcgtt ttgggtaact 600 agaggtgaga aaaggattca ttcctgttgt ccaaatctag atattcggga tgatcagaga 660 gaaatttctc gacagatatt tcttactgct attggtgatc aagctagaag tggtaagaga 720 cagatgtcgg agaatgagct gtggatgtat gaccaatttc gtgaaaatat tgctgcgcct 780 aacgcggtta ggtgcaataa tacatatcag ggttgtacat gtaggggttt ttctgatggt 840 aagaagaaag gcgcgcagta tgcgatagct cttcacagcc tgtatgactt caagttgaaa 900 gacttgatgg ctactatggt tgagaagaaa actaaagtgg ttcatgctgc tatgcttttt 960 gctcctgaaa gtatgttagt ggacgaaggt ccattacctt ctgttgacgg ttactacatg 1020 aagaagaacg ggaagatcta tttcggtttt gagaaagatc cttccttttc ttacattcat 1080 gactgggaag agtacaagaa gtatctactg gggaagccag tgagttacca agggaatgtg 1140 ttctacttcg aaccgtggca ggtgagagga gacacaatgc ttttttcgat ctacaggata 1200 gctggagttc cgaggaggtc tctatcatcg caagagtact accgaagaat atatatcagt 1260 agatgggaaa gcatggttgt tgtcccaatt ttcgatctgg tcgaatcaac gcgagagttg 1320 gtcaagaaag acctgtttgt agagaaacaa ttcatggaca agtgtttgga ttacatagct 1380 aggttatctg accagcagct gaccataagc aatgttaaat catacttgag ttcaaataat 1440 tgggtcttat tcataaacgg ggcggccgtg aagaacaagc aaagtgtaga ttctcgagat 1500 ttacagttgt tggctcaaac tttgctagtg aaggaacaag tggcgagacc tgtcatgagg 1560 gagttgcgtg aagcaattct gactgagacg aaacctatca cgtcattgac tgatgtgctg 1620 ggtttaatat caagaaaact gtggaagcag tttgctaaca agatcgcagt cggcggattc 1680 gttggcatgg ttggtactct aattggattc tatccaaaga aggtactaac ctgggcgaag 1740 gacacaccaa atggtccaga actatgttac gagaactcgc acaaaaccaa ggtgatagta 1800 tttctgagtg ttgtgtatgc cattggagga atcacgctta tgcgtcgaga catccgagat 1860 ggactggtga aaaaactatg tgatatgttt gatatcaaac ggggggccca tgtcttagac 1920 gttgagaatc cgtgccgcta ttatgaaatc aacgatttct ttagcagtct gtattcggca 1980 tctgagtccg gtgagaccgt tttaccagat ttatccgagg taaaagccaa gtctgataag 2040 ctattgcagc agaagaaaga aatcgctgac gagtttctaa gtgcaaaatt ctctaactat 2100 tctggcagtt cggtgagaac ttctccacca tcggtggtcg gttcatctcg aagcggactg 2160 ggtctgttgt tggaagacag taacgtgctg acccaagcta gagttggagt ttcaagaaag 2220 gtagacgatg aggagatcat ggagcagttt ctgagtggtc ttattgacac tgaagcagaa 2280 attgacgagg ttgtttcagc cttttcagct gaatgtgaaa gaggggaaac aagcggtaca 2340 aaggtgttgt gtaaaccttt aacgccacca ggatttgaga acgtgttgcc agctgtcaaa 2400 cctttggtca gcaaaggaaa aacggtcaaa cgtgtcgatt acttccaagt gatgggaggt 2460 gagagattac caaaaaggcc ggttgtcagt ggagacgatt ctgtggacgc tagaagagag 2520 tttctgtact acttagatgc ggagagagtc gctcaaaatg atgaaattat gtctctgtat 2580 cgtgactatt cgagaggagt tattcgaact ggaggtcaga attacccgca cggactggga 2640 gtgtgggatg tggagatgaa gaactggtgc atacgtccag tggtcactga acatgcttat 2700 gtgttccaac cagacaaacg tatggatgat tggtcgggat acttagaagt ggctgtttgg 2760 gaacgaggta tgttggtcaa cgacttcgcg gtcgaaagga tgagtgatta tgtcatagtt 2820 tgcgatcaga cgtatctttg caataacagg taataatcct ctctcttgat atttttaaat 2880 tatagaatta attagtttac tttattcttt actatatgat ttaaatagtt taatcttgtt 2940 tttgagtaaa ctattcgatt ttgatatttg tattcgtcct acaaagttgg aaatactgat 3000 gatattttct tttgaacgtg atacctacca atactaatct tacggaatct tttaatagag 3060 cactaatcaa catggaacta aagaccaatt cttaagtgtc tctgttgtac agttcatttt 3120 agtagtgcgt ttaagtatta ttatctccct tcatgcgggg caattatgta gattaaaatc 3180 gaaattatat aaaatttaca taagtctaag tctagggtct ccagctaatt gttatttttt 3240 taacgatgtt gactaaagca ataacgacgt tgacttgtgt taaacaggtt gatcttggac 3300 aatttaagtg ccctggatct aggaccagtt aactgttctt ttgaattagt tgacggtgta 3360 cctggttgtg gtaagtcgac aatgattgtc aactcagcta atccttgtgt cgatgtggtt 3420 ctctctactg ggagagcagc aaccgacgac ttgatcgaga gattcgcgag caaaggtttt 3480 ccatgcaaat tgaaaaggag agtgaagacg gttgattctt ttttgatgca ttgtgtcgat 3540 ggttctttaa ccggagacgt gttgcatttc gacgaagctc tcatggccca tgctggtatg 3600 gtgtactttt gcgctcagat agctggtgct aaacgatgta tctgtcaagg agatcagaat 3660 caaatttctt tcaagcctag ggtatctcaa gttgatttga ggttttctag tctggtcgga 3720 aagtttgaca ttgttacaga aaaaagagaa acttacagaa gtccagcaga tgtggctgcc 3780 gtattgaaca agtactatac tggagatgtc agaacacata acgcgactgc taattcgatg 3840 acggtgagga agattgtgtc taaagaacag gtttctttga agcctggtgc tcagtacata 3900 actttccttc agtctgagaa gaaggagttg gtaaatttgt tggcattgag gaaagtggca 3960 gctaaagtga gtacagtaca cgagtcgcaa ggagagacat tcaaagatgt agtcctagtc 4020 aggacgaaac ctacggatga ctcaatcgct agaggtcggg agtacttaat cgtggcattg 4080 tcgcgtcaca cacaatcact tgtgtatgaa actgtgaaag aggacgatgt aagcaaagag 4140 atcagggaaa gtgccgcgct tacgaaggcg gctttggcaa gattttttgt tactgagacc 4200 gtcttatgac ggtttcggtc taggtttgat gtctttagac atcatgaagg gccttgcgcc 4260 gttccagatt caggtacgat tacggacttg gagatgtggt acgacgcttt gtttccggga 4320 aattcgttaa gagactcaag cctagacggg tatttggtgg caacgactga ttgcaatttg 4380 cgattagaca atgttacgat caaaagtgga aactggaaag acaagtttgc tgaaaaagaa 4440 acgtttctga aaccggttat tcgtactgct atgcctgaca aaaggaagac tactcagttg 4500 gagagtttgt tagcattgca gaaaaggaac caagcggcac ccgatctaca agaaaatgtg 4560 cacgcgacag ttctaatcga agagacgatg aagaagctga aatctgttgt ctacgatgtg 4620 ggaaaaattc gggctgatcc tattgtcaat agagctcaaa tggagagatg gtggagaaat 4680 caaagcacag cggtacaggc taaggtagta gcagatgtga gagagttaca tgaaatagac 4740 tattcgtctt acatgtatat gatcaaatct gacgtgaaac ctaagactga tttaacaccg 4800 caatttgaat actcagctct acagactgtt gtgtatcacg agaagttgat caactcgttg 4860 ttcggtccaa ttttcaaaga aattaatgaa cgcaagttgg atgctatgca accacatttt 4920 gtgttcaaca cgagaatgac atcgagtgat ttaaacgatc gagtgaagtt cttaaatacg 4980 gaagcggctt acgactttgt tgagatagac atgtctaaat tcgacaagtc ggcaaatcgc 5040 ttccatttac aactgcagct ggagatttac aggttatttg ggctggatga gtgggcggcc 5100 ttcctttggg aggtgtcgca cactcaaact actgtgagag atattcaaaa tggtatgatg 5160 gcgcatattt ggtaccaaca aaagagtgga gatgctgata cttataatgc aaattcagat 5220 agaacactgt gtgcactctt gtctgaatta ccattggaga aagcagtcat ggttacatat 5280 ggaggagatg actcactgat tgcgtttcct agaggaacgc agtttgttga tccgtgtcca 5340 aagttggcta ctaagtggaa tttcgagtgc aagattttta agtacgatgt cccaatgttt 5400 tgtgggaagt tcttgcttaa gacgtcatcg tgttacgagt tcgtgccaga tccggtaaaa 5460 gttctgacga agttggggaa aaagagtata aaggatgtgc aacatttagc cgagatctac 5520 atctcgctga atgattccaa tagagctctt gggaactaca tggtggtatc caaactgtcc 5580 gagtctgttt cagaccggta tttgtacaaa ggtgattctg ttcatgcgct ttgtgcgcta 5640 tggaagcata ttaagagttt tacagctctg tgtacattat tccgagacga aaacgataag 5700 gaattgaacc cggctaaggt tgattggaag aaggcacaga gagctgtgtc aaacttttac 5760 gactggtaat atggaagaca agtcattggt caccttgaag aagaagactg gcgcgccacg 5820 tgttaattaa ctgattcgac taggcgcctc aatgtggaag aactgaacag ttcggattac 5880 attgaaggcg attttaccga tcaagaggtt ttcggtgagt tcatgtcttt gaaacaagtg 5940 gagatgaaga cgattgaggc gaagtacgat ggtccttaca gaccagctac tactagacct 6000 aagtcattat tgtcaagtga agatgttaag agagcgtcta ataagaaaaa ctcgtcttaa 6060 tgcataaaga aatttattgt caatatgacg tgtgtactca agggttgtgt gaatgaagtg 6120 ttcctttcgg gattgatcgt ttgtttgctt tttgatttta ttttatattg ttatctgttt 6180 ctgtgtatag actgtttgag attggcgctt ggccgactca ttgtcttacc ataggggaac 6240 ggactttgtt tgtgttgtta ttttatttgt attttattaa aattctcaat gatctgaaaa 6300 ggcctcgagg ctaagagatt attggggggt gagtaagtac ttttaaagtg atgatggtta 6360 caaaggcaaa aggggtaaaa cccctcgcct acgtaagcgt tattacgccc g 6411 4 6358 DNA Artificial Sequence Description of Artificial Sequence pBTA-delta-REP-delta-MP 4 gatccataaa acatttcaat cctttgaacg cggtagaacg tgctaattgg attttggtga 60 gaacgcggta gaacgtactt atcacctaca gttttatttt gtttttcttt ttggtttaat 120 ctatccagct tagtaccgag tgggggaaag tgactggtgt gcctaaaacc ttttctttga 180 tactttgtaa aaatacatac agatacaatg gcgaacggta acttcaagtt gtctcaattg 240 ctcaatgtgg acgagatgtc tgctgagcag aggagtcatt tctttgactt gatgctgact 300 aaacctgatt gtgagatcgg gcaaatgatg caaagagttg ttgttgataa agtcgatgac 360 atgattagag aaagaaagac taaagatcca gtgattgttc atgaagttct ttctcagaag 420 gaacagaaca agttgatgga aatttatcct gaattcaata tcgtgtttaa agacgacaaa 480 aacatggttc atgggtttgc ggctgctgag cgaaaactac aagctttatt gcttttagat 540 agagttcctg ctctgcaaga ggtggatgac atcggtggtc aatggtcgtt ttgggtaact 600 agaggtgaga aaaggattca ttcctgttgt ccaaatctag atattcggga tgatcagaga 660 gaaatttctc gacagatatt tcttactgct attggtgatc aagctagaag tggtaagaga 720 cagatgtcgg agaatgagct gtggatgtat gaccaatttc gtgaaaatat tgctgcgcct 780 aacgcggtta ggtgcaataa tacatatcag ggttgtacat gtaggggttt ttctgatggt 840 aagaagaaag gcgcgcagta tgcgatagct cttcacagcc tgtatgactt caagttgaaa 900 gacttgatgg ctactatggt tgagaagaaa actaaagtgg ttcatgctgc tatgcttttt 960 gctcctgaaa gtatgttagt ggacgaaggt ccattacctt ctgttgacgg ttactacatg 1020 aagaagaacg ggaagatcta tttcggtttt gagaaagatc cttccttttc ttacattcat 1080 gactgggaag agtacaagaa gtatctactg gggaagccag tgagttacca agggaatgtg 1140 ttctacttcg aaccgtggca ggtgagagga gacacaatgc ttttttcgat ctacaggata 1200 gctggagttc cgaggaggtc tctatcatcg caagagtact accgaagaat atatatcagt 1260 agatgggaaa gcatggttgt tgtcccaatt ttcgatctgg tcgaatcaac gcgagagttg 1320 gtcaagaaag acctgtttgt agagaaacaa ttcatggaca agtgtttgga ttacatagct 1380 aggttatctg accagcagct gaccataagc aatgttaaat catacttgag ttcaaataat 1440 tgggtcttat tcataaacgg ggcggccgtg aagaacaagc aaagtgtaga ttctcgagat 1500 ttacagttgt tggctcaaac tttgctagtg aaggaacaag tggcgagacc tgtcatgagg 1560 gagttgcgtg aagcaattct gactgagacg aaacctatca cgtcattgac tgatgtgctg 1620 ggtttaatat caagaaaact gtggaagcag tttgctaaca agatcgcagt cggcggattc 1680 gttggcatgg ttggtactct aattggattc tatccaaaga aggtactaac ctgggcgaag 1740 gacacaccaa atggtccaga actatgttac gagaactcgc acaaaaccaa ggtgatagta 1800 tttctgagtg ttgtgtatgc cattggagga atcacgctta tgcgtcgaga catccgagat 1860 ggactggtga aaaaactatg tgatatgttt gatatcaaac ggggggccca tgtcttagac 1920 gttgagaatc cgtgccgcta ttatgaaatc aacgatttct ttagcagtct gtattcggca 1980 tctgagtccg gtgagaccgt tttaccagat ttatccgagg taaaagccaa gtctgataag 2040 ctattgcagc agaagaaaga aatcgctgac gagtttctaa gtgcaaaatt ctctaactat 2100 tctggcagtt cggtgagaac ttctccacca tcggtggtcg gttcatctcg aagcggactg 2160 ggtctgttgt tggaagacag taacgtgctg acccaagcta gagttggagt ttcaagaaag 2220 gtagacgatg aggagatcat ggagcagttt ctgagtggtc ttattgacac tgaagcagaa 2280 attgacgagg ttgtttcagc cttttcagct gaatgtgaaa gaggggaaac aagcggtaca 2340 aaggtgttgt gtaaaccttt aacgccacca ggatttgaga acgtgttgcc agctgtcaaa 2400 cctttggtca gcaaaggaaa aacggtcaaa cgtgtcgatt acttccaagt gatgggaggt 2460 gagagattac caaaaaggcc ggttgtcagt ggagacgatt ctgtggacgc tagaagagag 2520 tttctgtact acttagatgc ggagagagtc gctcaaaatg atgaaattat gtctctgtat 2580 cgtgactatt cgagaggagt tattcgaact ggaggtcaga attacccgca cggactggga 2640 gtgtgggatg tggagatgaa gaactggtgc atacgtccag tggtcactga acatgcttat 2700 gtgttccaac cagacaaacg tatggatgat tggtcgggat acttagaagt ggctgtttgg 2760 gaacgaggta tgttggtcaa cgacttcgcg gtcgaaagga tgagtgatta tgtcatagtt 2820 tgcgatcaga cgtatctttg caataacagg taataatcct ctctcttgat atttttaaat 2880 tatagaatta attagtttac tttattcttt actatatgat ttaactgttc ttttgaatta 2940 gttgacggtg tacctggttg tggtaagtcg acaatgattg tcaactcagc taatccttgt 3000 gtcgatgtgg ttctctctac tgggagagca gcaaccgacg acttgatcga gagattcgcg 3060 agcaaaggtt ttccatgcaa attgaaaagg agagtgaaga cggttgattc ttttttgatg 3120 cattgtgtcg atggttcttt aaccggagac gtgttgcatt tcgacgaagc tctcatggcc 3180 catgctggta tggtgtactt ttgcgctcag atagctggtg ctaaacgatg tatctgtcaa 3240 ggagatcaga atcaaatttc tttcaagcct agggtatctc aagttgattt gaggttttct 3300 agtctggtcg gaaagtttga cattgttaca gaaaaaagag aaacttacag aagtccagca 3360 gatgtggctg ccgtattgaa caagtactat actggagatg tcagaacaca taacgcgact 3420 gctaattcga tgacggtgag gaagattgtg tctaaagaac aggtttcttt gaagcctggt 3480 gctcagtaca taactttcct tcagtctgag aagaaggagt tggtaaattt gttggcattg 3540 aggaaagtgg cagctaaagt gagtacagta cacgagtcgc aaggagagac attcaaagat 3600 gtagtcctag tcaggacgaa acctacggat gactcaatcg ctagaggtcg ggagtactta 3660 atcgtggcat tgtcgcgtca cacacaatca cttgtgtatg aaactgtgaa agaggacgat 3720 gtaagcaaag agatcaggga aagtgccgcg cttacgaagg cggctttggc aagatttttt 3780 gttactgaga ccgtcttatg acggtttcgg tctaggtttg atgtctttag acatcatgaa 3840 gggccttgcg ccgttccaga ttcaggtacg attacggact tggagatgtg gtacgacgct 3900 ttgtttccgg gaaattcgtt aagagactca agcctagacg ggtatttggt ggcaacgact 3960 gattgcaatt tgcgattaga caatgttacg atcaaaagtg gaaactggaa agacaagttt 4020 gctgaaaaag aaacgtttct gaaaccggtt attcgtactg ctatgcctga caaaaggaag 4080 actactcagt tggagagttt gttagcattg cagaaaagga accaagcggc acccgatcta 4140 caagaaaatg tgcacgcgac agttctaatc gaagagacga tgaagaagct gaaatctgtt 4200 gtctacgatg tgggaaaaat tcgggctgat cctattgtca atagagctca aatggagaga 4260 tggtggagaa atcaaagcac agcggtacag gctaaggtag tagcagatgt gagagagtta 4320 catgaaatag actattcgtc ttacatgtat atgatcaaat ctgacgtgaa acctaagact 4380 gatttaacac cgcaatttga atactcagct ctacagactg ttgtgtatca cgagaagttg 4440 atcaactcgt tgttcggtcc aattttcaaa gaaattaatg aacgcaagtt ggatgctatg 4500 caaccacatt ttgtgttcaa cacgagaatg acatcgagtg atttaaacga tcgagtgaag 4560 ttcttaaata cggaagcggc ttacgacttt gttgagatag acatgtctaa attcgacaag 4620 tcggcaaatc gcttccattt acaactgcag ctggagattt acaggttatt tgggctggat 4680 gagtgggcgg ccttcctttg ggaggtgtcg cacactcaaa ctactgtgag agatattcaa 4740 aatggtatga tggcgcatat ttggtaccaa caaaagagtg gagatgctga tacttataat 4800 gcaaattcag atagaacact gtgtgcactc ttgtctgaat taccattgga gaaagcagtc 4860 atggttacat atggaggaga tgactcactg attgcgtttc ctagaggaac gcagtttgtt 4920 gatccgtgtc caaagttggc tactaagtgg aatttcgagt gcaagatttt taagtacgat 4980 gtcccaatgt tttgtgggaa gttcttgctt aagacgtcat cgtgttacga gttcgtgcca 5040 gatccggtaa aagttctgac gaagttgggg aaaaagagta taaaggatgt gcaacattta 5100 gccgagatct acatctcgct gaatgattcc aatagagctc ttgggaacta catggtggta 5160 tccaaactgt ccgagtctgt ttcagaccgg tatttgtaca aaggtgattc tgttcatgcg 5220 ctttgtgcgc tatggaagca tattaagagt tttacagctc tgtgtacatt attccgagac 5280 gaaaacgata aggaattgaa cccggctaag gttgattgga agaaggcaca gagagctgtg 5340 tcaaactttt acgactggta atatggaaga caagtcattg gtcaccttga agaagaagac 5400 tggcgcgcca cgtgttaatt aactgattcg actaggcgcc tcaatgtgga agaactgaac 5460 agttcggatt acattgaagg cgattttacc gatcaagagg ttttcggtga gttcatgtct 5520 ttgaaacaag tggagatgaa gacgattgag gcgaagtacg atggtcctta cagaccagct 5580 actactagac ctaagtcatt attgtcaagt gaagatgtta agagagcgtc taataagaaa 5640 aactcgtctt aatgcataaa gaaatttatt gtcaatatga cgtgtgtact caagggttgt 5700 gtgaatgaag tcactgttct tggtcacgag acgtgtagta tcggtcatgc taacaaattg 5760 cgaaagcaag ttgctgacat ggttggtgtc acacgtaggt gtgcggaaaa taattgtgga 5820 tggtttgtct gtgttgttat caatgatttt acttttgatg tgtataattg ttgtggccgt 5880 agtcaccttg aaaagtgtcg taaacgtgtt gaaacaagaa atcgagaaat ttggaaacaa 5940 attcgacgaa atcaagctga aaacatgtct gcgacagcta aaaagtctca taattcgaag 6000 acctctaaga agaaattcaa agaggacaga gaatttggga caccaaaaag atttttaaga 6060 gatgatgttc ctttcgggat tgatcgtttg tttgcttttt gattttattt tatattgtta 6120 tctgtttctg tgtatagact gtttgagatt ggcgcttggc cgactcattg tcttaccata 6180 ggggaacgga ctttgtttgt gttgttattt tatttgtatt ttattaaaat tctcaatgat 6240 ctgaaaaggc ctcgaggcta agagattatt ggggggtgag taagtacttt taaagtgatg 6300 atggttacaa aggcaaaagg ggtaaaaccc ctcgcctacg taagcgttat tacgcccg 6358 5 6003 DNA Artificial Sequence Description of Artificial Sequence pBTA-delta-REP-delta-MP-delta-16K 5 gatccataaa acatttcaat cctttgaacg cggtagaacg tgctaattgg attttggtga 60 gaacgcggta gaacgtactt atcacctaca gttttatttt gtttttcttt ttggtttaat 120 ctatccagct tagtaccgag tgggggaaag tgactggtgt gcctaaaacc ttttctttga 180 tactttgtaa aaatacatac agatacaatg gcgaacggta acttcaagtt gtctcaattg 240 ctcaatgtgg acgagatgtc tgctgagcag aggagtcatt tctttgactt gatgctgact 300 aaacctgatt gtgagatcgg gcaaatgatg caaagagttg ttgttgataa agtcgatgac 360 atgattagag aaagaaagac taaagatcca gtgattgttc atgaagttct ttctcagaag 420 gaacagaaca agttgatgga aatttatcct gaattcaata tcgtgtttaa agacgacaaa 480 aacatggttc atgggtttgc ggctgctgag cgaaaactac aagctttatt gcttttagat 540 agagttcctg ctctgcaaga ggtggatgac atcggtggtc aatggtcgtt ttgggtaact 600 agaggtgaga aaaggattca ttcctgttgt ccaaatctag atattcggga tgatcagaga 660 gaaatttctc gacagatatt tcttactgct attggtgatc aagctagaag tggtaagaga 720 cagatgtcgg agaatgagct gtggatgtat gaccaatttc gtgaaaatat tgctgcgcct 780 aacgcggtta ggtgcaataa tacatatcag ggttgtacat gtaggggttt ttctgatggt 840 aagaagaaag gcgcgcagta tgcgatagct cttcacagcc tgtatgactt caagttgaaa 900 gacttgatgg ctactatggt tgagaagaaa actaaagtgg ttcatgctgc tatgcttttt 960 gctcctgaaa gtatgttagt ggacgaaggt ccattacctt ctgttgacgg ttactacatg 1020 aagaagaacg ggaagatcta tttcggtttt gagaaagatc cttccttttc ttacattcat 1080 gactgggaag agtacaagaa gtatctactg gggaagccag tgagttacca agggaatgtg 1140 ttctacttcg aaccgtggca ggtgagagga gacacaatgc ttttttcgat ctacaggata 1200 gctggagttc cgaggaggtc tctatcatcg caagagtact accgaagaat atatatcagt 1260 agatgggaaa gcatggttgt tgtcccaatt ttcgatctgg tcgaatcaac gcgagagttg 1320 gtcaagaaag acctgtttgt agagaaacaa ttcatggaca agtgtttgga ttacatagct 1380 aggttatctg accagcagct gaccataagc aatgttaaat catacttgag ttcaaataat 1440 tgggtcttat tcataaacgg ggcggccgtg aagaacaagc aaagtgtaga ttctcgagat 1500 ttacagttgt tggctcaaac tttgctagtg aaggaacaag tggcgagacc tgtcatgagg 1560 gagttgcgtg aagcaattct gactgagacg aaacctatca cgtcattgac tgatgtgctg 1620 ggtttaatat caagaaaact gtggaagcag tttgctaaca agatcgcagt cggcggattc 1680 gttggcatgg ttggtactct aattggattc tatccaaaga aggtactaac ctgggcgaag 1740 gacacaccaa atggtccaga actatgttac gagaactcgc acaaaaccaa ggtgatagta 1800 tttctgagtg ttgtgtatgc cattggagga atcacgctta tgcgtcgaga catccgagat 1860 ggactggtga aaaaactatg tgatatgttt gatatcaaac ggggggccca tgtcttagac 1920 gttgagaatc cgtgccgcta ttatgaaatc aacgatttct ttagcagtct gtattcggca 1980 tctgagtccg gtgagaccgt tttaccagat ttatccgagg taaaagccaa gtctgataag 2040 ctattgcagc agaagaaaga aatcgctgac gagtttctaa gtgcaaaatt ctctaactat 2100 tctggcagtt cggtgagaac ttctccacca tcggtggtcg gttcatctcg aagcggactg 2160 ggtctgttgt tggaagacag taacgtgctg acccaagcta gagttggagt ttcaagaaag 2220 gtagacgatg aggagatcat ggagcagttt ctgagtggtc ttattgacac tgaagcagaa 2280 attgacgagg ttgtttcagc cttttcagct gaatgtgaaa gaggggaaac aagcggtaca 2340 aaggtgttgt gtaaaccttt aacgccacca ggatttgaga acgtgttgcc agctgtcaaa 2400 cctttggtca gcaaaggaaa aacggtcaaa cgtgtcgatt acttccaagt gatgggaggt 2460 gagagattac caaaaaggcc ggttgtcagt ggagacgatt ctgtggacgc tagaagagag 2520 tttctgtact acttagatgc ggagagagtc gctcaaaatg atgaaattat gtctctgtat 2580 cgtgactatt cgagaggagt tattcgaact ggaggtcaga attacccgca cggactggga 2640 gtgtgggatg tggagatgaa gaactggtgc atacgtccag tggtcactga acatgcttat 2700 gtgttccaac cagacaaacg tatggatgat tggtcgggat acttagaagt ggctgtttgg 2760 gaacgaggta tgttggtcaa cgacttcgcg gtcgaaagga tgagtgatta tgtcatagtt 2820 tgcgatcaga cgtatctttg caataacagg taataatcct ctctcttgat atttttaaat 2880 tatagaatta attagtttac tttattcttt actatatgat ttaactgttc ttttgaatta 2940 gttgacggtg tacctggttg tggtaagtcg acaatgattg tcaactcagc taatccttgt 3000 gtcgatgtgg ttctctctac tgggagagca gcaaccgacg acttgatcga gagattcgcg 3060 agcaaaggtt ttccatgcaa attgaaaagg agagtgaaga cggttgattc ttttttgatg 3120 cattgtgtcg atggttcttt aaccggagac gtgttgcatt tcgacgaagc tctcatggcc 3180 catgctggta tggtgtactt ttgcgctcag atagctggtg ctaaacgatg tatctgtcaa 3240 ggagatcaga atcaaatttc tttcaagcct agggtatctc aagttgattt gaggttttct 3300 agtctggtcg gaaagtttga cattgttaca gaaaaaagag aaacttacag aagtccagca 3360 gatgtggctg ccgtattgaa caagtactat actggagatg tcagaacaca taacgcgact 3420 gctaattcga tgacggtgag gaagattgtg tctaaagaac aggtttcttt gaagcctggt 3480 gctcagtaca taactttcct tcagtctgag aagaaggagt tggtaaattt gttggcattg 3540 aggaaagtgg cagctaaagt gagtacagta cacgagtcgc aaggagagac attcaaagat 3600 gtagtcctag tcaggacgaa acctacggat gactcaatcg ctagaggtcg ggagtactta 3660 atcgtggcat tgtcgcgtca cacacaatca cttgtgtatg aaactgtgaa agaggacgat 3720 gtaagcaaag agatcaggga aagtgccgcg cttacgaagg cggctttggc aagatttttt 3780 gttactgaga ccgtcttatg acggtttcgg tctaggtttg atgtctttag acatcatgaa 3840 gggccttgcg ccgttccaga ttcaggtacg attacggact tggagatgtg gtacgacgct 3900 ttgtttccgg gaaattcgtt aagagactca agcctagacg ggtatttggt ggcaacgact 3960 gattgcaatt tgcgattaga caatgttacg atcaaaagtg gaaactggaa agacaagttt 4020 gctgaaaaag aaacgtttct gaaaccggtt attcgtactg ctatgcctga caaaaggaag 4080 actactcagt tggagagttt gttagcattg cagaaaagga accaagcggc acccgatcta 4140 caagaaaatg tgcacgcgac agttctaatc gaagagacga tgaagaagct gaaatctgtt 4200 gtctacgatg tgggaaaaat tcgggctgat cctattgtca atagagctca aatggagaga 4260 tggtggagaa atcaaagcac agcggtacag gctaaggtag tagcagatgt gagagagtta 4320 catgaaatag actattcgtc ttacatgtat atgatcaaat ctgacgtgaa acctaagact 4380 gatttaacac cgcaatttga atactcagct ctacagactg ttgtgtatca cgagaagttg 4440 atcaactcgt tgttcggtcc aattttcaaa gaaattaatg aacgcaagtt ggatgctatg 4500 caaccacatt ttgtgttcaa cacgagaatg acatcgagtg atttaaacga tcgagtgaag 4560 ttcttaaata cggaagcggc ttacgacttt gttgagatag acatgtctaa attcgacaag 4620 tcggcaaatc gcttccattt acaactgcag ctggagattt acaggttatt tgggctggat 4680 gagtgggcgg ccttcctttg ggaggtgtcg cacactcaaa ctactgtgag agatattcaa 4740 aatggtatga tggcgcatat ttggtaccaa caaaagagtg gagatgctga tacttataat 4800 gcaaattcag atagaacact gtgtgcactc ttgtctgaat taccattgga gaaagcagtc 4860 atggttacat atggaggaga tgactcactg attgcgtttc ctagaggaac gcagtttgtt 4920 gatccgtgtc caaagttggc tactaagtgg aatttcgagt gcaagatttt taagtacgat 4980 gtcccaatgt tttgtgggaa gttcttgctt aagacgtcat cgtgttacga gttcgtgcca 5040 gatccggtaa aagttctgac gaagttgggg aaaaagagta taaaggatgt gcaacattta 5100 gccgagatct acatctcgct gaatgattcc aatagagctc ttgggaacta catggtggta 5160 tccaaactgt ccgagtctgt ttcagaccgg tatttgtaca aaggtgattc tgttcatgcg 5220 ctttgtgcgc tatggaagca tattaagagt tttacagctc tgtgtacatt attccgagac 5280 gaaaacgata aggaattgaa cccggctaag gttgattgga agaaggcaca gagagctgtg 5340 tcaaactttt acgactggta atatggaaga caagtcattg gtcaccttga agaagaagac 5400 tggcgcgcca cgtgttaatt aactgattcg actaggcgcc tcaatgtgga agaactgaac 5460 agttcggatt acattgaagg cgattttacc gatcaagagg ttttcggtga gttcatgtct 5520 ttgaaacaag tggagatgaa gacgattgag gcgaagtacg atggtcctta cagaccagct 5580 actactagac ctaagtcatt attgtcaagt gaagatgtta agagagcgtc taataagaaa 5640 aactcgtctt aatgcataaa gaaatttatt gtcaatatga cgtgtgtact caagggttgt 5700 gtgaatgaag tgttcctttc gggattgatc gtttgtttgc tttttgattt tattttatat 5760 tgttatctgt ttctgtgtat agactgtttg agattggcgc ttggccgact cattgtctta 5820 ccatagggga acggactttg tttgtgttgt tattttattt gtattttatt aaaattctca 5880 atgatctgaa aaggcctcga ggctaagaga ttattggggg gtgagtaagt acttttaaag 5940 tgatgatggt tacaaaggca aaaggggtaa aacccctcgc ctacgtaagc gttattacgc 6000 ccg 6003 6 945 DNA Artificial Sequence Description of Artificial Sequence A. thaliana partial cDNA sequence sulphur gene 6 ccttcactct cttctccttc ctcaaaacct tcctcctccc ccatttgctt caggccaggt 60 aaattgtttg gaagcaagtt aaatgcagga atccaaataa ggccaaagaa gaacaggtct 120 cgttaccatg tttcggttat gaatgtagcc actgaaatca actctactga acaagtagta 180 gggaagtttg attcaaagaa gagtgcgaga ccggtttatc catttgcagc tatagtaggg 240 caagatgaga tgaagttatg tcttttgttg aatgttattg atccaaagat tggtggtgtt 300 atgattatgg gagatagagg aactggaaaa tctacaactg ttagatcatt agttgatctg 360 ttacctgaga ttaatgtagt tgcaggtgac ccgtataact cggatccgat agatcctgag 420 tttatgggtg ttgaagtaag agagagagtt gagaaaggag agcaagttcc tgttattgcg 480 actaagatta atatggttga tcttcctttg ggtgcaacag aagatagagt ttgtggaacc 540 atcgatatcg aaaaggcttt gacagaaggt gtaaaagcct ttgagcctgg tttgttggct 600 aaagctaata gagggattct ttatgttgat gaagttaatc tcttggatga tcatttggtt 660 gatgttcttt tggattcagc tgcttctggt tggaatacgg ttgagagaga agggatttcg 720 atttctcacc cggcgaggtt tatcttgatc ggttcaggaa atccggaaga aggagagctt 780 aggccacagc ttcttgatcg gtttggtatg catgcacaag tagggacggt tagagatgct 840 gatttacggg tcaagattgt tgaagagaga gctcgtttcg atagtaaccc aaaggatttc 900 cgtgacactt acaaaaccga gcaggacaag cttcaagacc agatt 945 7 469 DNA Artificial Sequence Description of Artificial Sequence A. thaliana partial cDNA sequence RUBISCO small subunit gene 7 cctctatgct ctcttccgct actatggttg cctctccggc tcaggccact atggtcgctc 60 ctttcaacgg acttaagtcc tccgctgcct tcccagccac ccgcaaggct aacaacgaca 120 ttacttccat cacaagcaac ggcggaagag ttaactgcat gcaggtgtgg cctccgattg 180 gaaagaagaa gtttgagact ctctcttacc ttcctgacct taccgattcc gaattggcta 240 aggaagttga ctaccttatc cgcaacaagt ggattccttg tgttgaattc gagttggagc 300 acggatttgt gtaccgtgag cacggtaact cacccggata ctatgatgga cggtactgga 360 caatgtggaa gcttcccttg ttcggttgca ccgactccgc tcaagtgttg aaggaagtgg 420 aagagtgcaa gaaggagtac cccaatgcct tcattaggat catcggatt 469 8 940 DNA Artificial Sequence Description of Artificial Sequence A. thaliana partial cDNA sequence LEAFY gene 8 ccatacggta tacgtttcta cacggcggcg aagatagcgg agttaggttt tacggcgagc 60 acgcttgtgg gtatgaagga cgaggagctt gaagagatga tgaatagtct ctctcatatc 120 tttcgttggg agcttcttgt tggtgaacgg tacggtatca aagctgccgt tagagctgaa 180 cggagacgat tgcaagaaga ggaggaagag gaatcttcta gacgccgtca tttgctactc 240 tccgccgctg gtgattccgg tactcatcac gctcttgatg ctctctccca agaagtgatt 300 ggacagggtt atctgaggaa ccggtgcagc aacaagacca gactgatgcg gcggggaata 360 acggcggagg aggaagtggt tactgggacg caggtcaagg aaagatgaag aagcaacagc 420 agcagagacg gagaaagaaa ccaatgctga cgtcagtgga aaccgacgaa gacgtcaacg 480 aaggtgagga tgacgacggg atggataacg gcaacggagg tagtggtttg gggacagaga 540 gacagaggga gcatccgttt atcgtaacgg agcctgggga agtggcacgt ggcaaaaaga 600 acggcttaga ttatctgttc cacttgtacg aacaatgccg tgagttcctt cttcaggtcc 660 agacaattgc taaagaccgt ggcgaaaaat gccccaccaa ggtgacgaac caagtattca 720 ggtacgcgaa gaaatcagga gcgagttaca taaacaagcc taaaatgcga cactacgttc 780 actgttacgc tctccactgc ctagacgaag aagcttcaaa tgctctcaga agagcgttta 840 aagaacgcgg tgagaacgtt ggctcatggc gtcaggcttg ttacaagcca cttgtgaaca 900 tcgcttgtcg tcatggctgg gatatagacg ccgtctttaa 940 9 793 DNA Artificial Sequence Description of Artificial Sequence mGFP5 cDNA sequence 9 aatgaagact aatctttttc tctttctcat cttttcactt ctcctatcat tatcctcggc 60 cgaattcagt aaaggagaag aacttttcac tggagttgtc ccaattcttg ttgaattaga 120 tggtgatgtt aatgggcaca aattttctgt cagtggagag ggtgaaggtg atgcaacata 180 cggaaaactt acccttaaat ttatttgcac tactggaaaa ctacctgttc catggccaac 240 acttgtcact actttctctt atggtgttca atgcttttca agatacccag atcatatgaa 300 gcggcacgac ttcttcaaga gcgccatgcc tgagggatac gtgcaggaga ggaccatctt 360 cttcaaggac gacgggaact acaagacacg tgctgaagtc aagtttgagg gagacaccct 420 cgtcaacagg atcgagctta agggaatcga tttcaaggag gacggaaaca tcctcggcca 480 caagttggaa tacaactaca actcccacaa cgtatacatc atggccgaca agcagaagaa 540 cggcatcaaa gccaacttca agacccgcca caacatcgaa gacggcggcg tgcaactcgc 600 tgatcattat caacaaaata ctccaattgg cgatggccct gtccttttac cagacaacca 660 ttacctgtcc acacaatctg ccctttcgaa agatcccaac gaaaagagag accacatggt 720 ccttcttgag tttgtaacag ctgctgggat tacacatggc atggatgaac tatacaaaca 780 tgacgaactc taa 793 10 29 DNA Artificial Sequence Description of Artificial Sequence Primer 10 ggggggatcc gggcgtaata acgcttacg 29 11 31 DNA Artificial Sequence Description of Artificial Sequence Primer 11 ggggggatcc ataaaacatt tcaatccttt g 31 12 22 DNA Artificial Sequence Description of Artificial Sequence Primer 12 ttagcaccag ctatctgagc gc 22 13 23 DNA Artificial Sequence Description of Artificial Sequence Primer 13 gttccaacca gacaaacgta tgg 23 14 43 DNA Artificial Sequence Description of Artificial Sequence Primer 14 cgtatctttg caataacagg taataatcct ctctcttgat att 43 15 42 DNA Artificial Sequence Description of Artificial Sequence Primer 15 ttaaattgtc caagatcaac ctgtttaaca caagtcaacg tc 42 16 22 DNA Artificial Sequence Description of Artificial Sequence Primer 16 tcgcacaaaa ccaaggtgat ag 22 17 33 DNA Artificial Sequence Description of Artificial Sequence Primer 17 ggattattac ctgttattgc aaagatacgt ctg 33 18 34 DNA Artificial Sequence Description of Artificial Sequence Primer 18 tgttaaacag gttgatcttg gacaatttaa gtgc 34 19 22 DNA Artificial Sequence Description of Artificial Sequence Primer 19 ttctcaaatc taggggccat tg 22 20 33 DNA Artificial Sequence Description of Artificial Sequence Primer 20 ccgaaaggaa cacttcattc acacaaccct tga 33 21 31 DNA Artificial Sequence Description of Artificial Sequence Primer 21 gaatgaagtg ttcctttcgg gattgatcgt t 31 22 22 DNA Artificial Sequence Description of Artificial Sequence Primer 22 actcactgat tgcgtttcct ag 22 23 43 DNA Artificial Sequence Description of Artificial Sequence Primer 23 ttaattaaca cgtggcgcgc cagtcttctt cttcaaggtg acc 43 24 41 DNA Artificial Sequence Description of Artificial Sequence Primer 24 ggcgcgccac gtgttaatta actgattcga ctaggcgcct c 41 25 31 DNA Artificial Sequence Description of Artificial Sequence Primer 25 ccttggcgcg ccttcactct cttctccttc c 31 26 32 DNA Artificial Sequence Description of Artificial Sequence Primer 26 ccccttaatt aatctggtct tgaagcttgt cc 32 27 31 DNA Artificial Sequence Description of Artificial Sequence Primer 27 ccttggcgcg cctctatgct ctcttccgct a 31 28 33 DNA Artificial Sequence Description of Artificial Sequence Primer 28 ccccttaatt aatccgatga tcctaatgaa ggc 33 29 33 DNA Artificial Sequence Description of Artificial Sequence Primer 29 ccttggcgcg ccatacggta tacgtttcta cac 33 30 30 DNA Artificial Sequence Description of Artificial Sequence Primer 30 ccccttaatt aaagacggcg tctatatccc 30 31 34 DNA Artificial Sequence Description of Artificial Sequence Primer 31 ggttggcgcg ccaatgaaga ctaatctttt tctc 34 32 33 DNA Artificial Sequence Description of Artificial Sequence Primer 32 ggggttaatt aattagagtt cgtcatgttt gta 33 

1. An insolated DNA vector which comprises: (a) transfer nucleotide sequence comprising (i) a plant active promoter, operably linked to (ii) a recombinant tobacco rattle virus (TRV) nucleic acid which includes: a sequence encoding a TRV trans acting factor, and cis acting elements, which confer on the TRV nucleic acid transcript the ability to replicate in the cytoplasm of a plant cell; and a heterologous nucleotide sequence which is foreign to said virus; (b) border sequences which permit the transfer of the transfer nucleotide sequence into a plant cell genome.
 2. A vector as claimed in claim 1 wherein the promoter is constitutive.
 3. A vector as claimed in claim 1 wherein the promoter is: developmentally regulated, inducible, or tissue specific.
 4. A vector as claimed in any one of the preceding claims comprising a transcriptional terminator within the transfer nucleotide sequence.
 5. A vector as claimed in any one of the preceding claims wherein the recombinant TRV nucleic acid does not encode a TRV coat protein.
 6. A vector as claimed in any one of the preceding claims wherein the recombinant TRV nucleic acid corresponds to all or part of TRV RNA
 1. 7. A vector as claimed in any one of the preceding claims wherein the recombinant TRV nucleic acid lacks all or part of the viral genome not required for replication in the cytoplasm.
 8. A vector as claimed in claim 6 which does not comprises an MP and\or 16K ORF.
 9. A vector as claimed in claim 7 or claim 8 wherein the heterologous nucleotide sequence is replaces part of the viral genome.
 10. A vector as claimed in any one of claims 1 to 8 wherein the heterologous nucleotide sequence is additional to the viral genome.
 11. A vector as claimed in any one of the preceding claims wherein the heterologous nucleotide sequence does not contain and is operably linked to a subgenomic promoter.
 12. A vector as claimed in any one of the preceding claims wherein the heterologous gene sequence is a multiple cloning site.
 13. A vector as claimed in claim 12 wherein the multiple cloning site comprises at least AscI, PmlI and PacI restriction enzyme sites.
 14. A vector as claimed in any one of claims 1 to 11 wherein the heterologous gene sequence is a targeting sequence which is capable of down-regulating expression of a target gene.
 15. A vector as claimed in claim 14 wherein the targeting sequence is, in a sense or anti-sense orientation, either: (a) complementary to a sequence within the target gene, or (b) homologous to a sequence within the target gene.
 16. A vector as claimed in claim 14 or claim 15 wherein the targeting sequence targets a conserved sequence within a target gene group such as to down-regulate expression of one or more members of a target gene group.
 17. A vector as claimed in any one claims 14 to 16 comprising more than one targeting sequence.
 18. A vector as claimed in any one of claims 14 to 17 wherein the target gene is: an endogenous plant gene, a transgene, or a gene from a pathogen.
 19. A vector as claimed in claim 18 wherein the endogenous plant gene is one associated with one or more of the following traits: ripening, pollen formation, lignin biosynthesis, flower pigment production, regulatory pathways controlling development, environmental responses, growth, disease resistance, toxin production.
 20. A vector as claimed in any one of the preceding claims which is an Agrobacterium binary vector.
 21. A vector as claimed in claim 20 which is derived from transformation vector pBINTRA6 (SEQ ID NO: 1).
 22. A vector as claimed in claim 21 which is pBTAΔMPΔ16K (SEQ ID NO: 3) or pBTAΔMP (SEQ ID NO: 2)
 23. A vector as claimed in any one of the preceding claims which is suitable for stable transformation of a plant cell.
 24. A host cell containing a vector of any one of the preceding claims.
 25. A method which includes the step of causing or allowing transcription from a vector of any one of claims 14 to
 23. within a plant cell to produce a cytoplasmically-replicating RNA.
 26. A method of silencing a target gene in to at least part of a plant, the method including: (i) introducing a vector of any one of claims 14 to 23 into at least part of a plant, (ii) causing or allowing transcription from the vector to produce a cytoplasmically-replicating RNA.
 27. A method as claimed in claim 26 wherein the vector is introduced by Agrobacterium-mediated transient transformation.
 28. A method of characterizing a target gene comprising the steps of: (a) silencing the target gene in at least part of a plant according to a method of claim 26 or claim 27, (b) observing the phenotype of the part of the plant in which the target gene has been silenced.
 29. A method as claimed in claim 28 wherein the phenotype is compared with a plant or part of a plant wherein the target gene is being expressed.
 30. A method as claimed in any one of claims 26 to 29 wherein the at least part of the plant includes the meristems of the plant.
 31. Use of a vector of any one of claims 14 to 23 in the production of a transgenic plant.
 32. A method of silencing a target gene in a plant, the method including: (i) introducing a vector of any one of claims 14 to 23 into a plant cell such that the transfer nucleotide sequence is stably incorporated into the genome of the cell, (ii) regenerating a plant from the plant cell.
 33. A plant cell having incorporated into its genome the transfer nucleotide sequence of any one of claims 14 to
 23. 34. A plant obtainable by the process of claim 32 comprising the plant cell of claim
 33. 35. A plant which is clone or descendant of the plant of claim 34 and which comprises the plant cell of claim
 33. 36. A plant as claimed in claim 34 or claim 35 which is selected from: Arabidopsis thaliana; Allium cepa; Amaranthus caudatus; Amaranthus retroflexus; Antirrhinum majus; snap-dragon; Arachis hypogaea; Avena sativa; Bellis perennis; Beta vulgaris; Brassica campestris; Brassica campestris ssp. napus; Brassica campestris ssp. pekinensis; Brassica juncea; Calendula officinalis; Capsella bursa-pastoris; Capsicum annuum; Catharanthus roseus; Cheiranthus cheiri; Chenopodium album; Chenopodium amaranticolor; Chenopodium foetidum; Chenopodium quinoa; Coriandrum sativum; Cucumis melo; Cucumis sativus; Glycine max; Gomphrena globosa; Gypsophila elegans; Helianthus annuus; Hyacinthus; Hyoscyamus niger; Lactuca sativa; Lathyrus odoratus; Linum usitatissimum; Lobelia erinus; Lupinus mutabilis; Lycopersicon esculentum; Lycopersicon pimpinellifolium; Melilotus albus; Momordica balsamina; Myosotis sylvatica; Narcissus pseudonarcissus; Nicandra physalodes; Nicotiana benthamiana; Nicotiana clevelandii; Nicotiana glutinosa; Nicotiana rustica; Nicotiana sylvestris; Nicotiana tabacum; Nicotiana edwardsonii; Ocimum basilicum; Petunia hybrida; Phaseolus vulgaris; Phytolacca americana; Pisum sativum; Raphanus sativus; Ricinus communis; Salvia splendens; Senecio vulgaris; Solanum melongena; Solanum nigrum; Solanum tuberosum; Spinacia oleracea; Stellaria media; Trifolium pratense; Trifolium repens; Tropaeolum majus; Tulipa; Vicia faba; Vicia villosa; Viola arvensis.
 37. A plant propagule of the plant of any of claims 34 to 36 and which comprises the plant cell of claim
 33. 38. An extract or derivative of the plant or plant propagule of any one of claims 34 to 37 and which comprises the plant cell of claim
 33. 39. A process for producing a vector of claim of any one of claims 14 to 23, which process comprises the step of cloning a targeting sequence into a vector of claim 12 or
 13. 